Amino acid derivatives inhibiting extracellular matrix metalloproteinase and TNF alpha release

ABSTRACT

The invention concerns compounds of general formula (X) in which Y represents in particular —CONHOH, R 1  represents in particular a C 1 -C 5  alkyl group, AA represents an amino acid, or an amino acid sequence, and R 3  represents in particular a group of formula —NH—(CH 2 ) 2 —SCH 3 . The invention also concerns the pharmaceutical compositions containing them, and the methods for obtaining them.

This application is a 371 of PCT/FR98/00801 filed Apr. 21, 1998.

This invention relates to new amino acid derivatives possessing aninhibiting action on metalloproteinase of the extracellular matrix, andmore particularly gelatinase, a process for the production of thesederivatives and pharmaceutical compositions containing them. Thesederivatives also possess an inhibiting action on the release of αTNF(Tumor Necrosis Factor) as well as on the production of αTGF (TumorGrowth Factor).

The breakdown of the extracellular matrix is due principally to theenzymatic action of metalloproteinase (MMP).

The enzymatic activity of this metalloproteinase is regulatedphysiologically by natural inhibitors such as TIMP (Tissue Inhibitor ofMetalloproteinase) or alpha-2-macroglobulin. An imbalance in theproduction of the enzymes and their inhibitors leads to a high proteinactivity observed in pathologies involving a process of breakdown of theextracellular matrix.

Compounds having the property of inhibiting the action of themetalloproteinase involved in the breakdown of the extracellular matrix,such as collagenase, gelatinase and stromelysine therefore may be usedin the treatment of pathologies in which metalloproteinase is involved,such as rheumatoid arthritis, osteoarthritis, osteoporosis, cornealulceration, periodontitis, gingivitis or tumorous invasion andmetastatic proliferation, atherosclerosis, AIDS, chronic inflammatorydiseases of the intestine, these examples not being restrictive.

αTNF is a pro-inflammatory cytokin which is produced initially in theform of an inactive 28 kDa precursor. The cleavage of this precursorleads to the release of an active form of 17 kDa involved in numerousinflammatory, immunological, infectious or malignant pathologies.Compounds inhibiting the release of αTNF therefore may be used in thetreatment of numerous pathologies in which αTNF is involved, such asrheumatoid arthritis, Crohn's disease, plaque sclerosis, septic shock,cancer or cachexia associated with an immunodeficiency, these examplesnot being restrictive.

αTGF is a growth factor forming part of the EGF (Epidermal GrowthFactor) family. It is produced by the embryonic tissues, keratinocytes,macrophages, eosinophiles, epitheliums (mammary gland and cornea),pancreas, gastric mucous membrane, pituitary and brain.

αTGF interacts with the EGF receptor; a cascade of reactions, resultingin mitosis, ensues. αTGF also is mitogenic for tumorous cells.

αTGF induces the transformation and growth of cells in vitro.

An overproduction of αTGF is observed in tumors as well as in cell stockderived from mammary tumors. αTGF also is involved in angiogenesis. Itlikewise stimulates hypercalcemia and inhibits gastric acid secretion.Finally, it is involved in inflammatory reactions.

Compounds inhibiting the production of αTGF therefore may be used in thetreatment of pathologies in which αTGF is involved, such as cancer,psoriasis, eczema, the formation of keloids, diabetic retinopathy,atherosclerosis, inflammatory diseases, these examples not beingrestrictive.

Numerous inhibitors of MMP and/or of TNF release already are known, themost active being the derivatives of hydroxamic acid with the generalformula I:

wherein R₁ represents an alkyl chain, generally isobutyl, and AA anamino acid or an amino acid chain. Such compounds are described, forexample, in patent applications EP 0214639, WO 93/20047, WO 94/02447, WO94/21625, WO 94/10990, WO 95/06031. MMP inhibitors inhibiting theproduction of αTGF are described in international application WO96/25156.

Other compounds have been described as inhibitors of matrixmetalloproteinase in which the hydroxamic function has been replaced bya thiol (general formula II) or phosphinic (general formula III)function.

Finally, derivatives of N-carboxymethyl peptides also have been claimed(general formula IV).

In these different families, the R₁ residue interacts with the subsiteS′₁ of the various enzymes. The stereochemistry of the carbon bearingthis residue is essential for activity and must be of preciseconfiguration, R, in the case of hydroxamic (J. Enzyme Inhibition,(1987), 2, 1-22) and phosphinic derivatives. None of the existingpatents describing the MMP inhibitors makes reference to disubstitutionon the R₁-bearing carbon.

The substitution of this carbon therefore is of extreme importance foractivity, and it has been shown in particular that substitution of thehydrogen in this carbon with a methyl remainder brings about a loss ofactivity of a factor of 300 between compound V and compound VI (J. Am.Chem. Soc. (1995), 117, 4671-4682).

R. P. ROBINSON et al. (Bioorg. Med. Chem. Letters (1996), 6, 1719-1724)also have studied the substitution of this carbon. The gemdisubstitution leads systematically to a significant loss of activity(compound VII versus compounds VIII and IX).

This invention is derived from the discovery made by the Inventors that,in a completely unexpected manner considering the state of the art setforth above, the compounds of the following general formula X:

characterized by a gem disubstitution of this carbon with an R₁ residueand an alcohol function are powerful inhibitors of metalloproteinase ofthe extracellular matrix, and more particularly of gelatinase(inhibiting concentration at 50%: C150<200 nM). These compounds alsoinhibit the release of αTNF from macrophages in mice stimulated by LPS(lipopolysaccharides) (C150: 10 to 0.01 μM) as well as the production ofαTGF.

This invention has as its purpose to provide new compounds inhibitingmetalloproteinase, and/or the release of αTNF and/or the production ofαTGF, the different inhibition activities of

R₆ represents —H, or a C₁ to C₆ alkoxy group, or a benzyloxy group,

R₇ represents —H, or a halogen atom such as —Cl or —Br,

R₁ represents:

a C₃ to C₁₆ linear or branched, or C₃ to C₆ cyclized alkyl chain, saidchain comprising, as the case may be, a heteroatom such as O, S or N,

a phenoxyalkyl or phenylalkyl group, substituted or unsubstituted, or aheteroarylalkyl group, the alkyl group being C₂ to C₅,

R₂ represents:

a hydrogen atom, or,

a C₁ to C₅ alkyl or C₂ to C₅ alkylidene group, or

a hydroxyl, a C₁ to C₆ alkoxy or a benzyloxy, provided that Y represents—CONHOH when R₂ represents a hydroxyl, or

a hydroxymethyl, or C₁ to C₆ alkoxymethyl group, or

an arylalkyl group in which the alkyl portion is C₁ to C₆, anaryloxymethyl group, an arylthiomethyl group, a heteroarylthiomethylgroup, in which aryl designates a phenyl remainder, possiblysubstituted, in particular by —OH, —OCH₃, a linear or branched C₁ to C₃alkyl group, a halogen such as —Cl or —Br, an amine group such as —NH₂,—NHCOCH₃, —NHCOOR₁₀, R₁₀ representing a linear or branched C₁ to C₃alkyl group, or

A phthalimide alkyl group in which the alkyl portion is C₁ to C₆, or

an alkoxycarbonylmethyl group (alkoxy designating methoxy, ethoxy), abenzyloxycarbonylmethyl, an acetylmethyl, provided that Y represents —SHin these three cases.

AA represents an amino acid, or an amino acid chain, these amino acidsbeing natural or otherwise, and advantageously with an absolute Sconfiguration, in particular an amino acid with the formula thesecompounds being comparable or even superior to those of compounds of thestate of the art described above.

This invention further has as its purpose to provide new medicinescontaining the aforementioned new compounds as active principle, andoffering the advantage of possessing a better bioavailability than thecompounds of the state of the art described above.

This invention likewise has as its purpose the use of the aforementionednew compounds for the preparation of new medicines described above,capable of being used in the context of treatment of pathologies inwhich the metalloproteinase of the extracellular matrix and/or αTNF areinvolved, as well as pathologies in which an overproduction of αTGF isinvolved.

This invention has as its subject compounds of the following generalformula (X):

in which:

Y represents:

—CONHOH, or

—SH, or

a group with the formula

 or

a group with the formula

in which:

R₄ represents —H, or a C₁ to C₆ alkyl group, or a phenylalkyl group inwhich the alkyl group is C₁ to C₆,

R₅ represents a group with the formula

in which R represents:

a C₁ to C₄ linear or branched alkyl chain,

a —CH₂—Y group in which Y represents a ring of 4 to 6 carbon atoms inthe ring comprising, as the case may be, one or several heteroatoms suchas O, S or N, said ring being aromatic or nonaromatic, substituted asthe case may be, in particular by one or several —OCH₃, —NO₂, —NH₂groups, or by one or several halogen atoms chosen in particular fromamong —Cl, —Br, —F and —I,

a group with the formula

R₃ represents a group with the formula —NH—(R₈)_(n)—R₉ in which:

n represents 0 or 1,

R₈ represents a linear or branched alkyl chain, with 1 to 8 carbon atomscomprising, as the case may be, one or several heteroatoms such as O orS,

R₉ represents a hydrogen atom or a methyl, nitrile, morpholino, phenyl,methoxy, hydroxyl, thiomethyl group, or a group with the formula—CH(NH₂)═N—OH, or a —N(CH₃)₂ group.

More particularly, the invention has as its subject the compoundscharacterized by the following general formula (Xa):

in which:

Y represents —CONHOH,

R₁ represents:

—CH(CH₃)₂,

More particularly, the invention has as its subject, by way of preferredcompounds, those a corresponding to the above-noted formulas X or Xa, inwhich R represents a group with the formula

in which R_(a) and R_(b) represent a halogen atom, in particular achlorine atom.

More particularly the invention also has as its subject, by way ofpreferred compounds, those corresponding to the above-noted formulas Xor Xa, in which R₃ represents a group with the formula

—NH —(CH₂)₂—SCH₃

The invention further has as its subject any mixture comprising, on theone hand, compounds with the following formula (XI.1):

in which Y, R₁ and R₂, AA and R₃ are such as defined above and, on theother, compounds with the following formula (XI.2):

in which Y, R₁ R₂, AA and R₃ have the meaning indicated hereinabove, theproportion of the compounds (XI.1) and XI.2) in the mixtureadvantageously being approximately 50% to approximately 99% for thecompound of formula (XI.1) and approximately 50% to approximately 1% forthe compound of formula (XI.2).

—CH₂—CH(CH₃)₂,

R₂ represents:

an alkyl group with 1 to 5 carbon atoms, in particular a methyl orpropyl group,

a hydroxyl, or

an alkoxy group with 1 to 5 carbon atoms, in particular a methoxy group,

R represents:

—C(CH₃)₃,

—CH₂—CH(CH₃)₂,

 aromatic or nonaromatic, in which R_(a) and R_(b), independently of oneanother, represent —H, —Cl, —Br, —I, —F, —OCH₃, —NO₂, —NH₂,

a group with the formula

R₃ represents a —NH—(CH₂)_(n1)—R₉ group in which:

n₁ represents 0, 1 or 2,

R₉ represents —CH₃, —C≡N, —COOCH₃, —SCH₃, —O—(CH₂)₂OH, —O—(CH₂)₂—OCH₃,—CH(NH₂)═N—OH,

Compounds particularly preferred in the context of this invention arethose possessing a stereochemistry such that the R₁ and R₂ substituentsare positioned in anti in relation to the succinic remainder inaccordance with the following formula (XI.1):

Compounds particularly preferred in the context of this invention arethose corresponding to the following formulas:

The invention likewise has as its subject any pharmaceutical compositioncomprising, as an active principle, a compound or compounds and/or amixture or mixtures such as described hereinabove, in combination withan acceptable pharmaceutical vehicle.

The pharmaceutical compositions according to the inventionadvantageously are presented in a form which may be administered orally,parenterally or rectally.

The pharmaceutical compositions according to the invention preferablyare characterized in that the dosage of active principle isapproximately 0.1 to approximately 500 mg/kg/day, preferably from 1 to300 mg/kg/day orally and rectally, and approximately 0.1 μg/kg/day to 1mg/kg/day parenterally.

Preferred pharmaceutical compositions according to the invention arepresented in a form which may be administered orally, in a unit dosageof 1 mg to 250 mg of active principle per dose, and preferably from 10mg to 250 mg of active principle per dose, at the rate of 1 to 4 dosesper day.

Pharmaceutical compositions also preferred according to the inventionare presented in a form which may be administered parenterally, in aunit dosage of 1 μg to 50 mg of active principle per injection, at therate of 1 to 2 injections per day.

The invention likewise has as its subject the use of a compound orcompounds and/or a mixture or mixtures such as described hereinabove,for the preparation of a medicine intended for the treatment of human oranimal pathologies in which metalloproteinase, and/or αTNF and/or αTGFare involved.

More particularly, the invention has as its subject the use of acompound or compounds and/or a mixture or mixtures such as describedhereinabove, for the preparation of a medicine having the property ofinhibiting the action of metalloproteinase involved in the breakdown ofthe extracellular matrix, such as the collagenase, gelatinase andstromelysine, this medicine being intended for the treatment of human oranimal pathologies linked to this action of metalloproteinase, inparticular for the treatment of:

rheumatoid arthritis,

osteoarthritis,

osteoporosis,

corneal ulceration,

periodontitis,

gingivitis,

tumorous invasions,

metastatic proliferation,

atherosclerosis,

AIDS,

chronic inflammatory diseases of the intestine,

neurodegenerative diseases such as Alzheimer's disease and plaquesclerosis.

More particularly the invention has as its subject the use of a compoundor compounds and/or a mixture or mixtures such as described hereinabove,for the preparation of a medicine having the property of inhibiting therelease of αTNF from its inactive precursor, this medicine beingintended for the treatment of human or animal pathologies in which αTNFis involved, in particular for the treatment of inflammatory,immunological, infectious or malignant pathologies, such as:

rheumatoid arthritis,

Crohn's disease,

plaque sclerosis,

septic shock,

cancer,

cachexia associated with an immunodeficiency.

More particularly the invention has as its subject the use of a compoundor compounds and/or a mixture or mixtures such as described hereinabove,for the preparation of a medicine having the property of inhibiting theproduction of αTGF, this medicine being intended for the treatment ofhuman or animal pathologies in which αTGF is involved, such as:

cancer,

psoriasis,

eczema,

formation of keloids,

diabetic retinopathy,

atherosclerosis,

inflammatory diseases.

Generally speaking, the various pathologies capable of being treated inthe context of this invention may be classified in the following manner:

I. Systemic inflammatory response syndrome, including:

septicemias, in particular Gram positive bacterium, Gram negativebacterium, fungal, or meningococcemia,

traumatisms and hemorrhages,

burns,

exposures to ionizer radiation,

acute pancreatitis,

respiratory distress syndrome in adults.

II. Reperfusion injuries, such as reperfusion ischemia.

III. Cardiovascular diseases, such as:

myocardial infarction,

congestive heart failure.

IV. Infectious diseases, including:

AIDS,

meningitis,

hepatitis,

arthritis,

periarthritis,

pneumonia,

epiglottitis,

0157:H7 E. Coli infection,

hemolytic uremic syndrome,

thrombolytic thrombocytopenic purpura,

malaria,

hemorrhagic dengue,

leishmaniasis,

leprosy,

septic shock,

streptococcal myositis,

gas gangrene,

tuberculosis,

orchitis,

legionnaires' disease,

lyme disease,

influenza,

infectious mononucleosis in Burkitt's lymphoma,

cancer of the rhinopharynx,

viral encephalitis.

V. In obstetrics, gynecology, including:

premature labor,

miscarriage,

sterility.

VI. Autoimmune inflammatory diseases, including:

rheumatoid arthritis and the seronegative arthropathies,

osteoarthritis,

Crohn's disease, ulcerative colitis,

lupus erythematosus,

iridocyclitis, uveitis and inflammation of the optic nerve,

idiopathic pulmonary fibrosis,

systemic vascularitis and Wegener's granulomatosis,

sarcoidosis,

orchitis.

VII. Allergic and atopic diseases, including:

asthma,

allergic rhinitis

eczema,

allergic contact dermatitis,

allergic conjunctivitis,

hypersensitivity pneumonitis.

VIII. Malignant diseases, including:

acute lymphoblastic leukemia,

acute monocytic leukemia,

chronic myeloid leukemia,

chronic lymphocytic leukemia,

Hodgkin's disease,

myeloid splenomegaly,

Kaposi's sarcoma,

colorectal carcinoma,

malignant histiocytosis,

paraneoplastic syndrome and hypercalcemia in malignant diseases,

IX. Transplants, including:

graft rejection

reaction of the graft against the host

X. Cachexia

XI. Congenital diseases, including:

mucoviscidosis,

familial lymphohistiocytosis,

sickle-cell anemia

XII. Dermatological diseases, including:

psoriasis,

alopecia.

XIII. Neurological diseases, including:

plaque sclerosis

headaches.

XIV. Kidney diseases, including:

nephritic syndrome,

hemodialysis,

uremia.

XV. Toxic treatments, including:

OKT3 therapy,

anti-CD3 therapy,

cytokine therapy,

chemotherapy,

radiotherapy,

chronic salicylate poisoning.

XVI. Idiopathic, metabolic diseases, including:

Wilson's disease,

hemochromatosis,

α1-antitrypsin deficiency,

diabetes,

Hashimoto's thyroiditis,

osteoporosis.

The invention also has as its subject the processes for preparation ofthe compounds or mixtures described hereinabove, and forming the subjectof the description which follows.

The abbreviations used in the description of the preparation processesof the invention and the detailed description of the experimentalportion hereinafter, are the following:

AcOEt Ethyl acetate α AD-mix Asymmetric dihydroxylation α mixture βAD-mix Asymmetric dihydroxylation β mixture Ar Aromatic Buli Butyllithium CH₂Cl₂ Dichloromethane DCC Dicyclohexyl carbodiimide DIPEADiisopropyl ethylamine DMF Dimethylformamide Et₂O Diethyl ether HMPTHexamethylphosphoretriamide HOBT Hydroxybenzotriazole LDA Lithiumdiisopropyl amidide LHMSA Lithium hexamethyldisyl amidide MeOH MethanolNa₂SO₄ Sodium sulfate NEt₃ Triethylamine PyBop Tris pyrrolidinobenzotriazolyl oxyphosphonium, hexaflurophosphate tBuOH Tert-butanol THFTetrahydrofuran THP Tetrahydropyran TMSCl Trimethylsilyl chloride TosParatoluenesulfonate WSC Water-soluble carbodiimide

The compounds according to the invention in which Y is a CONHOH group(also designated hereinafter as compounds of formula XV) may be obtainedaccording to the following Diagram I:

in which:

step 1 consists in condensing α-hydroxysuccinic acid XII (where R₈ is aprotective group compatible with the various elements of the moleculesuch as t-butyl or benzyl) with an AA-R₃ remainder where AA and R₃ aresuch as defined previously, by a method of coupling used in peptidesynthesis and preferably PyBop at room temperature for 1 to 24 hours (inthe case where R₂═OH, the alcohols may be protected beforehand with, forexample, a silyl derivative).

step 2 consists in hydrolyzing the ester XIII obtained in the precedingstep into carboxylic acid XIV with trifluroacetic acid, in particular atroom temperature in a solvent such as CH₂Cl₂ for 1 to 10 hours when R₈is t-butyl, or in hydrogenolyzing the ester XIII into acid XIV with, forexample, H₂ Pd/C when R₈ is benzyl (in particular under atmosphericpressure in a polar solvent such as ethanol for 30 minutes to 10 hours),

in step 3, hydroxamic acid XV is formed by reaction of hydroxylamine,protected O hydroxylamine or diprotected N,O hydroxylamine,preferentially with O—THP hydroxylamine or O-benzyl hydroxylamine (whenR₂ is other than alkylidene, aryloxymethyl and heteroarylthiomethyl) inthe presence of a coupling reagent such as DCC/HOBT or WSC/HOBT at roomtemperature in a solvent such as THF, CH₂Cl₂, or DMF for 1 to 24 hours(when R₂═OH, the alcohols are protected beforehand with, for example,TMSCl); the (di)protected O or N—O hydroxylamines then are deprotectedaccording to the nature of the protective group, for example in an acidmedium for O—THP hydroxylamine (in particular at room temperature in aTHF—H₂O mixture for 1 to 24 hours) or H₂ Pd/C for O-benzyl hydroxylamine(in particular under atmospheric pressure in a polar solvent such asethanol for 30 minutes to 10 h.).

The compounds XIV and XV also may be obtained when R₂ is other thanheteroarylthiomethyl and R₁ other than heteroarylalkyl through thereactions of the following Diagram 2:

in which:

steps 4 and 6 are performed as in steps 1 and 3 of diagram 1respectively, and starting from compounds XVI and XIV, which leads tothe compounds of formula XVII and XV respectively,

step 5 consists in oxidizing the double ethylene bond of the compound offormula XVII into acid, in particular by ozonolysis (for example at −60°C. in CH₂Cl₂ until obtaining a steady blue color), then oxidation (inparticular at room temperature with NaClO₂ and NaH₂PO₄ in tBuOH—H₂O for15 hours) or directly by KMnO₄/NalO₄ (in particular at room temperaturein a tBuOH—H₂O mixture for 1 to 10 hours), which leads to the compoundof formula XIV.

The compounds XIV and XV also may be prepared according to the sequenceof reactions (except R₂═OH, alkoxy or benzyloxy) of the followingDiagram 3:

in which:

steps 9 and 10 are identical to steps 2 and 3 of Diagram 1 and areperformed starting from compounds XIII and XIV respectively, which leadsto compounds XIV and XV respectively,

step 7 consists in reacting the sodium salt of a ketoacid XVIII withAA-R₃ by means of a coupling agent, for example oxalyl chloride with DMFat room temperature for 1 to 10 hours.

step 8 consists in a Reformatsky reaction (Rathke, Org. Reac. 22,423-460, 1975) between compound XIX and a bromo-ester with the formula:

 (in which R₂ and R₈ are the very same as before) in the presence ofzinc (in particular in a benzene-diethyl ether mixture, at 80° C. for 1hour 30 minutes); this reaction leads to a mixture of stereoisomerswhich may be separated, for example, by a chromatographic method toproduce compounds XIII.

The succinic acids XII may be prepared by Reformatsky reaction performedas previously according to the following Diagram 4 between, on the onehand, a compound with the formula R₂—CHBr—CO—OR₈ in which R₂ is such asdefined above (except R₂═OH, alkoxy, benzyloxy) and R₈ is such asdefined hereinbelow and, on the other, a compound of formula XX in whichR₁ is such as defined hereinabove, and R₉ is such as definedhereinbelow:

in which R₈ and R₉ are carboxylic acid protective groups which may becleaved selectively: R₉ may be, for example, a benzyl remaindersensitive to catalytic hydrogenolysis and R₈ a saponifiable ethyl ort-butyl group sensitive to acid hydrolysis.

Under these conditions, the reaction produces a mixture of the fourdiastereoisomers of formula XXI which may be separated, for example, bya chromatographic method to produce compounds XII).

The compounds XII also may be obtained through Evans oxazolidinones (J.Am. Chem. Soc. 104, 1737-1739, 1982; J. Am. Chem. Soc. 112, 8215-8216,1990) according to the sequence of reactions (except R₂═OH, alkoxy orbenzyloxy) of the following Diagram 5:

in which:

step 11 consists in acylating the oxazolidinone XXII previously treatedwith Buli (in particular at −70° C. in THF) or with NaH (in particularat 0° C. in THF) with an acid chloride

 (in particular at room temperature for 15 hours), in which R₂ is suchas defined above,

step 12 consists in condensing the enolate of the derivative XXII(prepared through the action of a base, for example LDA, LHMSA at −60°C. in THF for 30 minutes or a Lewis acid, for example TiCl₄ at 0° C. inCH₂Cl₂ for 1 hour) with a ketoester XX, in particular at −60° C. in THFor CH₂Cl₂ for 2 hours, R₉ being a carboxylic acid protective group(chiral or otherwise) compatible with the following step; under theseconditions, the reaction leads to a mixture of stereoisomers which maybe separated, for example, by a chromatographic method,

step 13 consists in cleaving the chiral copula of the compound XXIV withan aqueous base, for example aqueous KOH (2N) compatible with R₉ or withLiOOH (prepared from LiOH+H₂O₂) so as to obtain carboxylic acid XXV, inparticular in a THF—H₂O mixture at room temperature for 1 hour 30minutes.

step 14 consists in cleaving the copula of compound XXIV with an organicbase, for example MeOMgBr, LiOBr, in particular in THF at 0° C. for 1hour 30 minutes, compatible with R₉ so as to obtain the ester directly,

step 15 consists in protecting the carboxylic acid of compound XXV withan R₈ protective group; R₈ and R₉ must be able to be cleavedselectively, for example R₈=t-butyl (isobutene in CH₂Cl₂ in the presenceof a catalytic quantity of acid such as sulfuric acid at roomtemperature in a closed receptacle for 1 to 24 hours) and R₉=benzyl(K₂CO₃ in acetonitrile in the presence of a benzyl halogenide at 80° C.for 1 to 10 hours).

step 16 consists in hydrolyzing the R₉ protective group of the compoundXXVI either in a basic medium, for example aqueous NaOH or in an acidmedium, for example trifluroacetic acid, or in hydrogenolyzing, forexample H₂ Pd/C, depending on the R₉ structure, so as to obtain succinicacids in the same manner as before.

A method for obtaining the compounds of structure XVI consists inperforming an aldolization reaction from a keto-ester XXVII and analkene XXVIII (in particular in the presence of a Lewis acid such asSnCl₄ at −80° C. in a solvent such as CH₂Cl₂ for 5 minutes to 2 hours),or from a keto-acid (in the form of sodium salt or triethylamine) XXXand an alkene XXXI (in particular at room temperature between 1 and 10hours in a THF—H₂O mixture) according to the following Diagram 6:

in which:

R₁ and R₂ have the same meaning as in Diagram 2,

R₁₀ is a possibly branched C₁-C₁₂ alkyl, a benzyl or an optically purecompound such as mandelic acid esterified with a linear or branchedC₁-C₃ alkyl, or a benzyl,

R₁₁ is a linear or branched C₁-C₃ alkyl, or a chlorine,

R₁₂ is sodium or triethylamine,

R₁₃ is hydrogen, a linear or branched C₁-C₃ alkyl; R₁₃ also mayrepresent a chain forming a ring with the boron atom such as, forexample, di-isopropyltartrate,

the reactions are diastereoselective and lead to stereochemistryderivatives XVI if the double bond is of Z geometry for the compoundsXXVIII and E geometry for the compounds XXXI.

A more particularly preferred method consists in adopting as an R₁₀substituent an optically pure compound such as ethyl ester mandelicacid, which makes it possible to obtain optically pure compounds XVI.

The compounds XXIX and XVI also can make it possible to obtain acid XIIaccording to the following reaction Diagram 7:

in which:

step 17 consists in esterifying the compound XVI so as to obtain one ofthe compounds XXIX, for example a benzyl or mandelic ester with PyBop,

step 18 consists in oxidizing the double bond of the compound XXIX in amanner identical to step 5 of Diagram 2,

step 19 consists in protecting the carboxylic acid with an R₉ groupcompatible with the deprotection of the R₁₀ group, for example R₉ ist-butyl when R₁₀ is ethyl mandelate,

step 20 consists in deprotecting the acid bearing R₁₀ in a manneridentical to step 16 of Diagram 5, which leads to the obtaining of thecompound of formula XII.

The keto-acids or esters XX, XXVII, XXX, when they are not commerciallyavailable, may be prepared through the following reaction Diagram 8:

R₁₄ is a linear or branched C₁-C₃ alkyl,

R₁₅ is R₁ less one carbon,

R₉, R₁₀ and R₁₂ are as before,

step 21 consists in reacting the corresponding ester in the presence ofa base, for example tBuOK, diethyl oxalate (addition of diethyl oxalateto tBuOK, in diethyl ether, at t<10° C., then addition of the ester atroom temperature and stirring at this temperature for 15 hours),

step 22 consists in hydrolyzing the esters in heated condition in anacid medium, for example sulfuric acid 5N, then neutralizing with abase, for example soda, to obtain the product XXX (R₁₂=Na⁺),

the compound XXX (R₂=HN⁺Et₃) is easily obtained through treatment of thesodium salt with triethylamine hydrochlorate,

step 23 consists in esterifying the compound XXX (R₁₂=Na⁺) through theconventional methods of esterification, for example oxalyl chloride,DMF.

The silyl derivatives XXVIII with Z geometry may be obtained accordingto the following Diagram 9:

in which:

R₂ is such as described in Diagram 2,

R₁₁ is such as described above,

step 24 consists in performing an alkylation of an alkyne by means of abase, for example t-Buli (in particular in diethyl ether at −70° C.),with ClSi(R₁₁)₃ (in particular at −70° C. in diethyl ether, then at roomtemperature between 5 and 45 minutes),

step 25 consists in reducing the triple bond to a double bond through areducing agent such as hydrogen in the presence of a catalyst such asNickel acetate/NaBH4 in ethanol at atmospheric pressure for 2 hours.

Another method for obtaining the XXVIII derivative (R₂=CH₃) is describedhereinbelow in Diagram 10:

in which:

R₁₁ is such as described before,

step 26 consists in reacting an alkylsilane with butadiene in thepresence of triethylaluminum and Ni acetylacetonate in a closedreceptacle, in particular at 60° C. for 5 to 20 hours,

the compound obtained is an E+Z mixture,

step 27 consists in isomerizing the double bond by heating thendistilling the product.

The hydroxamic acids for which R₂=OH or alkoxy or benzyloxy may beobtained according to the following Diagram 11:

in which:

R₁, R₈, R₉ are such as described in Diagram 4 except when R₂=benzyloxy,R₈ and R₉ cannot be cleaved by hydrogenoiysis;

R₁₆ is linear or branched C₁-C₅ alkyls or represents a chain and forms aring with the two oxygen atoms,

step 28 consists in performing a Wittig reaction between the compoundsXX and a phosphonium salt with the structure:

 (X being a halogen) in DMF at room temperature for 1 to 10 hours or aphosphonate with the structure

R₁₇ being a linear or branched C₁-C₃ alkyl,

this reaction making it possible to lead to a mixture of E and Z alkeneswhich must be separated, for example by a chromatographic method, so asto obtain a compound XXXVI of E geometry,

step 29 consists in performing a Sharpless asymmetric hydroxylation(Chem. Rev. 2483-2547, 1994) in the presence of βAD-mix andmethanesulfonamide, in particular in a tBuOH—H₂O mixture at roomtemperature for 1 to 10 hours; this reaction leads to an optically pureproduct; the use of α or βAD-mix on alkene Z leads to two otherdiastereoisomers,

step 30 is identical to step 16 of Diagram 5,

step 31 consists in hydrolyzing the ester into carboxylic acid in abasic or acid medium depending on the structure of R₈,

step 32 consists in protecting the α-hydroxyacid in the form ofdioxolane by reacting it with an acetal, for example2,2-dimethoxypropane in DMF at 50° C., 15 hours,

step 33 consists in deprotecting the ester by hydrogenolysis; the R₉ inthis case is exclusively of benzyl type to be compatible with thedioxolane which does not tolerate the aqueous basic or acid medium,

step 34 consists in coupling the AA-R₃ amino acid by a method compatiblewith dioxolane, for example PyBop as before;

step 35 consists in substituting hydroxylamine for the dioxolane in anMeOH—H₂O mixture at −20° C. for 1 to 15 minutes,

steps 36, 37 and 38 are identical to steps 1, 2 and 3 of Diagram 1,

step 39 consists in alkylating the secondary alcohol by reacting it witha base, for example NaH, then an electrophile, for example an alkyl orbenzyl halogenide, in particular in THF at room temperature for 1 to 10hours,

step 40 is identical to step 16 of Diagram 5,

steps 41, 42 and 43 are identical to steps 1, 2 and 3 of Diagram 1.

The compounds according to the invention in which Y=SH,

may be obtained according to the following Diagram 12:

in which:

step 44 consists in opening an epoxide of formula L in which R₁ and R₂are such as defined hereinabove, and R₉ is a carboxylic acid protectivegroup, in particular a benzyl remainder sensitive to catalytichydrogenolysis, this opening of the epoxide L being accomplished with anucleophile, for example a thiol protected by an R₁₈ group compatiblewith R₉, for example a benzyl in methanol for 1 hour at 60° C.,

step 47 consists in deprotecting the ester LI, for example withtrifluroacetic acid as before,

step 48 is identical to step 1 of Diagram 1, and performed starting fromcompound LII obtained in the preceding step,

step 49 consists in deprotecting the sulfur, for example with sodium inliquid ammonia, in particular at −60° C. for 5 to 15 minutes,

step 45 consists in opening the epoxide L with protected hydroxylaminesuch as defined previously, for example R₁₉=benzyl or THP as previously,

step 50 consists in deprotecting the LV ester by a method compatiblewith R₁₉.

step 51 is identical to step 48 and performed starting from the compoundLVI obtained in the preceding step,

step 52 consists in reacting the hydroxylamine LVII with formic acid andacetic anhydride, in particular at a temperature of at least 100° C. for1 to 15 hours,

step 53 consists in cleaving R₁₉ on the compound LVIII with, forexample, H₂ Pd/C or HCl 1N depending on the structure of R₁₉ as before,

step 46 consists in opening an epoxide L with hypophosphorous acid withthe formula H₃PO₂, then esterification with a coupling agent such as DCCand an R₄OH group in which R₄ is such as defined hereinabove, in thepresence, for example, of trimethylorthoformate and tetramethylguanidineat room temperature for 5 hours,

step 54 consists in treating the compound LIX with a compound with theformula:

 (prepared according to the methods described in the literature) inwhich R₆ and R₇ are such as defined hereinabove, in particular in CH₂Cl₂in the presence of bis trimethylsilyl acetamide at room temperature for5 hours, which leads to the obtaining of the compound of formula LX inwhich R₅ represents:

step 55 consists in cleaving the R₉ ester by a method compatible with R₄as previously,

step 56 is identical to step 48, and performed starting from thecompound LXI obtained in the preceding step,

step 57 consists in cleaving the R₄ group of the compound LXII obtainedin the preceding step, for example with the aid of Nal in acetone underreflux for 15 hours.

The compounds of Diagram 12 are mixtures of diastereoisomers oroptically pure, depending on the original compound L. The mixtures ofdiastereoisomers may be separated, for example, by a chromatographicmethod.

Preparation of the compounds L in a racemic manner according to thefollowing Diagram 13:

in which:

R₁, R₂, R₉ are such as described previously,

step 58 consists in performing a Wittig reaction between a phosphoniumsalt

 (R₂ and X⁻ are the very same as before) and the compound XX in thepresence of a base, for example BuLi in THF at a temperature rangingbetween 0° C. and 60° C. for 1 hour; the olefin LXIV obtained is an Eand Z mixture which may be separated, for example, by a chromatographicmethod.

step 59 consists in oxidizing the double bond with, for example,KMnO₄-acetic acid in acetone at a temperature of −10° C. for 1 hour 30minutes.

step 60 consists in reducing the carbonyl with a reducing agent, forexample NaBH₄ in ethanol at 0° C. for 15 minutes,

step 61 consists in transforming the secondary alcohol into a startergroup by reacting it with, for example, methane-sulfonic acid in thepresence of a base, for example NEt₃ in diethyl ether at 0° C. for 1hour,

step 62 consists in treating the compound LXVIII with a base, forexample NAH, to form epoxide in DMF at room temperature for 1 to 3hours.

The olefin LXIV with E geometry also may be obtained through reactionsof the following Diagram 14:

in which:

step 63 consists in performing a chloration of the compound LXIX with,for example, sulfuryl chloride in dichloromethane at 35° C. for 30minutes.

step 64 consists in performing an alkylation in the presence of a base,for example NaH in a THF—HMPT mixture at room temperature for 15 hours,

step 65 consists in carrying out a dealcoxycarbonylation followed by anelimination with, for example, LiCl by heating in a solvent such as DMF,DMSO or HMPT. The product obtained under these conditions has Egeometry.

Preparation of optically pure epoxide according to the following diagram15:

in which:

step 64 consists in performing an asymmetric dihydroxylation as beforewith the aid of βAD-mix to lead to the compound LXXIV; the enantiomermay be obtained by αAD-mix and the other two diastereoisomers startingfrom E olefin and β or αAD-mix.

step 67 is identical to steps 61 and 62 of Diagram 13.

Experimental Portion

Intermediary 1: 2(S*) hydroxy-2 (S*)-(3-methylpropyl) 3 (R*) methylpent-4-enoic acid

Method A

a) (E)-but-2-enyl boronic bis (methyl-2-propyl ester) acid

To 23.8 g (212 mmoles) of tBuOK in 175 ml of dry THF cooled to −78° C.,add 21 ml (230 mmoles) of trans-2-butene. Then add in 1 hour 30 minutes,98 ml (212 mmoles) of nBuli (2.45 M in hexane). At the end of addition,stir for ½ hour at −50° C. Cool to −78° C. and add 49 ml (212 mmoles) oftriisopropyl borate, stir for ½ hour.

Add 200 ml of N HCl saturate with NaCl. Extract with 4 times 200 ml ofethyl ether. Collect the ethereal phases, dry over sodium sulfate.

Add 35.8 ml (467 mmoles) of isopropyl alcohol, 66 g of anhydrous sodiumsulfate and stir for 1 night at room temperature.

Decant the inorganic, evaporate the ether at 30° C. in the rotaryevaporator (under vacuum). Recover 17.95 g of oil or 45%. To bepreserved under nitrogen: Eb: 30° C./0.3 mmHg.

RMN (CDCl₃): δ5.5 (m, 2H, CH+CH); 4.4 (m, 2H, OCH—(Me)₂); 1.7 (m, 5H, CH₃—CH═ and ═CH—CH ₂—B; 1.3 (m, 12H, O—CH—(CH ₃)₂).

b) 2(S*)hydroxy-2(S*) 3-methylpropyl 3 (R*) methyl pent-4-enoic acid

Disperse 3.77 g (24.8 mmoles) of 4-methyl-2-oxo-sodium pentanoate in 30ml of CH₂Cl₂. Add 30 ml of HCl N saturated with NaCl. Extract 3×15 ml ofCH₂Cl₂. Collect and dry the organic phases over Na₂SO₄. Filter andintroduce this solution into a 250-ml three-necked flask.

Cool at −25° C., add 3.48 ml (24.8 mmoles) of triethlamine, then add4.57 g (24.8 mmoles) of product a) and stir for one night at roomtemperature. Pour over HCl 6N, extract with CH₂Cl₂, dry over Na₂So₄.Filter. Evaporate.

Purify by flash chromatography on 200 g of silica (eluant: CH₂Cl₂:MeOH;95:5).

Recover 2.73 g of white solid (yield 60%).

P.F.: 86° C.

IR (CDCl₃): v CO: 1706 cm⁻¹; v C═C: 1639 cm⁻¹

RMN (CDCl₃): δ5.8 (m,1H, CH₂═CH); 5.15 (m, 2H, CH ₂═CH—); 2.5 (m, 1H,═CH—CH(CH₃)—); 1.75 (m, 3H, CH ₂—CH—(CH₃)₂); 1 (3d, 9H, CH ₃).

Method B

To 4.8 g (42.7 mmoles) of tBuOK in 35 ml of THF at −78° C., add 4.2 ml(45.2 mmoles) of trans-2-butene. Without exceeding −65° C., add 21.35 ml(42.7 mmoles) of nBuli 2 M in hexane in 1 hour. At the end of addition,stir for ½ hour at −50°C., then cool again to −78° C. and add 9.85 ml(42.7 mmoles) of triisopropyl borate and stir for 30 minutes at −78° C.

Solubilize 6.49 g (42.7 mmoles) of 4-methyl-2-sodium oxovalerate in 15ml of water. Add this solution to the reaction medium and stir for onenight at room temperature.

Acidify with HCl 6N and extract with 3 times 50 ml of ethyl acetate.

Purify by flash chromatography on 600 g of silica (eluant:Ch₂Cl₂:MeOH:AcOH, 97:3:0.3). Recover 6.05 g of product b) or 77%. RMNidentical to b) of method A.

Intermediary 2: (Z) but-2-enyl triethylsilane

In a mini-autoclave cooled to −20° C., introduce:

9.8 ml (113 mmoles) of butadiene

18 ml (113 mmoles) of triethylsilane

60 mg (0.22 mmoles) of nickel acetylacetonate

0.32 ml (2.3 mmoles) of triethylaluminum.

Stir for 24 hours at 60° C. Distill at 50-53° C. under 7 mmHg. Recover12.7 g (67%) of product Z. RMN (CDCl₃): δ5.4 (2H, m,—CH═CH); 1.6 (3H, m,CH ₃—CH═); 1.55 (2H, m, HC═CH—CH ₂); 1 (9H, m, 3CH ₃); 0.5 (6H, m, 3CH₃Si).

Intermediary 3: 1(Z) hex-2-enyl triethylsilane

a) 1(hex-3-ynyl) triethylsilane

Under nitrogen atmosphere, introduce 8.8. ml (13 mmoles) of tBuli 1.45 Min pentane. Add Et₂O (12 ml), 1.84 ml (12 mmoles) of TMEDA, 1.37 ml (12mmoles) of 2-hexane and allow to return to 0° C.

Stir for 1 hour at 0° C., then cool again to −78° C. and introduce 2.45ml (15 mmoles) of chlorotriethylsilane. Allow to return to +20° C. inapproximately 45 minutes.

Add 20 ml of water. Extract with diethyl ether. Dry. Evaporate. Distillat 75% under 0.4 mmHg in the ball kiln. Recover 2.76 g (or 100%) ofproduct.

RMN (CDCl₃): δ2.15 (2H, m,—CH ₂—C≡); 1.5 (4H, m, CH ₂—CH₂—C≡CH—CH ₂—Si);1 (12H, m, Si(CH₂—CH ₃)₃ and CH ₃—CH₂—CH₂); 0.65 (6H, m, Si(CH ₂—CH₃)₃).

b) 1-(Z)hex-2-enyl triethylsilane

To 9.5 ml of absolute ethanol containing 0.5 ml of soda 2N, add 400 mgof NaBH₄. Stir for 10 minutes. Filter in 15 ml of absolute ethanolcontaining 370 mg (1.5 mmoles) of Nickel acetate, add 1.5 ml (1.5mmoles) of the filtered solution. Place under hydrogen atmosphere. Add2.35 g (12 mmoles) of product a) and stir for 2 hours at roomtemperature. Filter on celite. Concentrate. Distill at 125° C./22 mmHg.Recover 1.64 g or 68%.

RMN (CDCl₃): δ5.4 and 5.25 (2 m, 2H, CH═CH); 2 (2H, m, CH ₂—CH═); 1.55(2H, d, ═CH—CH ₂—Si); 1.4 (m, 2H, CH₃—CH ₂—CH₂—CH═); 1 (12H, m, CH₃—CH₂—CH₂ and Si(CH₂—CH ₃)₃); 0.55 (6H, m, Si(CH ₂—CH₃)₃).

Intermediary 4: 2(S)[1(S) (1,1 dimethyl) ethoxycarbonyl) ethyl]2(S)hydroxy-4-methyl pentanoic acid

a) 4-Methyl-2-sodium oxopentanoate

In a 20-l reactor, introduce 771.8 g (6 moles) of tBuOK and 6 l of ethylether. Purge with nitrogen. Cool to +8° C. Add in 1 hour 846 ml (6moles) of ethyl oxalate. Return to +20° C. and add in 20 minutes 900 ml(6 moles) of ethyl isovalerate and stir at room temperature for onenight. Freeze at 0° C. and add 6 l of HCl N. Extract with ethyl ether.Dry (Na₂SO₄) and evaporate the organic phases.

Take up the oil obtained with 3 l of dioxane and 3 l of H₂SO₄ 5N andheat for 4 days at 100° C. Freeze and neutralize with 2.2 l of NaOH 10N(pH 7). Wash 2 times with AcOEt. Evaporate the aqueous phase to dryness.Dry the solid obtained in the vane pump. Take up with 7 l of methanol.Mix, filter, concentrate. Recrystallize the 1,020 g obtained in 6.4 l ofabsolute ethanol. Recover 454 g (50%) of pure product.

RMN (CD₃OD): δ2.6 (2H, d, CH—CH ₂COCOONa); 2.15 (1H, m, (CH₃)₂—CH—CH₂);0.95 (6H, d, (CH ₃)₂ CH).

b) (S) ethyl mandelate

In 20 ml of dry CH₂Cl₂, add 1.52 g (10 mmoles) of mandelic acid S,εDMAP, 1.70 ml (21 mmoles) of pyridine and εDMF. Add 2.7 ml (21 mmoles)of TMSCl and stir for 2 hours at room temperature. Add 0.91 ml (10.5mmoles) of oxalyl chloride and stir for 2 hours at room temperature. Add20 ml of ethanol and stir for ½ hour at room temperature. Wash with 2×20ml of HCl N then NaHCO₃. Dry (Na₂SO₄), evaporate to dryness. Recover1.73 g (96%).

[α]_(D)=127.7° at t=21° C. (c=3, CHCl₃).

RMN (CDCl₃): δ7.4 (5H, m, CH(Ar)); 5,2 (1H, s, Ar—CH—OH); 4.2 (2H, m,OCH ₂); 1.25 (3H, t, OCH₂CH ₃).

c) 4-methyl-2-oxo pentanoic 1 (S) ethoxycarbonyl phenyl methyl esteracid

Disperse 50 g (0,329 mole) of methyl-4-oxo-2 sodium pentanoate in 900 mlof CH₂Cl₂ containing εDMF. Add dropwise 28.7 ml (0.329 mole) of oxalylchloride. At the end of addition, stir for 30 minutes at roomtemperature. Cool to +10° C. and add 56.4 g (0.313 M) of compound b)solubilized in 300 ml of CH₂Cl₂ Then add 57.3 ml (0.411 mole) oftriethylamine diluted in 200 ml of CH₂Cl₂. Stir for one night at roomtemperature. Wash with HCl N, then NaHCO₃. Dry, evaporate. Purify byflash chromatography on 800 g of silica (eluant: heptane:AcOEt; 95:5).Recover 71 g of oil (78%).

RMN (CDCl₃): δ7.5 (5H, m, H(Ar); 6 (1H, s,—O—CH—CO₂Et); 4.2 (2H, m, OCH₂—Ch₃); 2.8 (2H, d, —CH ₂—COCO); 2.3 (1H, m, CH—(CH₃)₂); 1.25 (3H, t,OCH₂—CH ₃); 1 (6H, d, (CH ₃)₂CH).

d) -2(S) hydroxy-2(S)[(2-methyl)propyl] 3(R)methyl pent-4-enoic 1 (S)ethoxy carbonyl phenylmethyl ester acid

Solubilize 10 g (34.2 mmoles) of compound c) in 200 ml of dry CH₂Cl₂.Cool to −78° C. and add 3.94 ml (34.2 mmoles) of SnCl₄. Stir for 30minutes at −78° C. and add 5.83 g (34.2 mmoles) of intermediary 2diluted in 50 ml of CH₂Cl₂. Stir for 1 hour 30 minutes at −78° C. andadd HCl N. Extract with CH₂Cl₂, dry (Na₂SO₄), evaporate. Eliminate thetriethylsilanol in the rotary evaporator (70° C. under 1 mmHg). Recover11.6 g of oil (97%).

RMN (CDCl₃): δ7.45 (5H, m, H(Ar)); 5.95 1H, s, OCHAr); 5.85 (1H, m,CH₂═CH); 5.1 (2H, m, CH ₂═); 4.25 (2H, m, OCH ₂—CH₃); 3.1 (1H, broad s,OH); 2.6 (1H, m, ═CH—CH(CH₃)—); 1.65 (3H, m, CH ₂—CH—(CH₃)₂₎; 1.3 (3H,t, OCH₂—CH ₃); 1.25 (3H, d, ═CH—CH(CH ₃)—); 0.95 (3H, d) and 0.7 (3H, d,—CH(CH ₃)₂)

e) 2 (S)[-1(-1(S) hydroxycarbonylethyl]-2 (S) hydroxy 4-methyl pentanoic(S) (ethoxycarbonyl) (phenyl)methyl ester acid

54.7 g (157 mmoles) of product d) are dispersed in 550 ml of dry CH₂Cl₂.Cool to −60° C. and ozonolyze to steady blue color. Purge with nitrogenand add 20.4 g (314 mmoles) of Zn and 18.3 ml (314 mmoles) of aceticacid. Stir for 1 hour at room temperature. Filter and evaporate. Take upthe residue obtained in 550 ml of tBuOH. Add 50 ml (471 mmoles) of2-methyl-2-butene. Then add an aqueous solution containing 48.9 g (314mmoles) of NaH₂PO₄, 2H₂O, 35.9 g (361 mmoles) of NaClO₂, H₂O 215 ml.

Stir for one night at room temperature. Add a saturated NaHCO₃ solution.Wash with pentane and extract with ethyl ether. Dry. Evaporate. Recover46.1 g of white oil (or 80%).

[α]_(D): +60.2° at t=20° C. (c=1, CHCl₃).

RMN (CDCl₃): δ7.45 (5H, s, HAr); 6 (1H, s, O—CH(CO)—Ar); 4.25 (2H, m OCH₂CH₃); 3.1 (1H, q, HO₂C—CH(CH₃)); 1.8 (3H, m, CH ₂—CH(CH₃)₂); 1.4 (3H,d, CH(CH ₃)—COOH); 1.3 (3H, t, OCH₂—CH ₃); 0.95 and 0.75 (6H, 2d, CH—CH₃)₂)

f) 2-(S) [1(S) (1,1 dimethyl ethoxycarbonyl)ethyl]-2(S) hydroxy-4-methylpentanoic (S) ethoxy carbonyl phenylmethyl ester acid

34.5 g (94.1 mmoles) of compound e) are solubilized in 330 ml of dryCH₂Cl₂. Cool the autoclave to −20° C., add 300 ml of isobutene, 0.4 mlof concentrated sulfuric acid. Close the autoclave and stir for onenight at room temperature. Pour over a saturated NaHCO₃ solution. Drythe organic phase, filter, evaporate to dryness. Recover 31.9 g of pureproduct (80%).

[α]_(D)=+70.2° at t=20° C. (c=1, MeOH)

RMN (CDCl₃): δ7.45 (5H, m, HAr); 6 (1H, s, O—CH Ar); 4.2 (2H, m, OCH₂CH₃); 3.75 (1H, broad s, OH); 2.9 (1H, q, tBuOCOCH); 1.7 (3H, m, CH₂CH(CH₃)₂); 1.5 (9H, s, (CH ₃)₃—C); 1.4 (3H, d, CH—CH ₃); 1.25 (3H, t,OCH₂CH ₃); 0.95 and 0.7 (6H, 2d, CH—CH ₃)₂).

g) 2-(S) [1 (S)((1,1 dimethyl) ethoxycarbonyl) ethyl] 2 (S) hydroxy4-methyl pentanoic acid

Solubilize 31.7 g (75 mmoles) of product f) in 320 ml of absoluteethanol. Under nitrogen, add 3.2 g of 10% Pd/C. Stir for 2 hours at 20°C. under hydrogen atmosphere. Filter the catalyst, rinse it in ethanol.Evaporate to dryness. Take up with ethyl ether, extract with 75 ml ofsoda N.

Wash the aqueous phase, then acidify with 75 ml of HCl N. Extract withethyl ether, dry, evaporate. Recover 18.7 g of white solid (95%).

M.P.: 67° C.

[α]_(D)=−9.9° C. at t=20° C. (c=1, CHCl₃)

RMN (CDCl₃): δ8.9 and 4.6 (2H very broad, OH and COOH); 2.75 (1H, q,tBuOCOCHCH₃); 1.85 (2H, m, CH ₂—CH); 1.55 (1H, m, CH ₂—CH); 1.5 (9H, s,(CH ₃)₃C); 1.2 (3H, d, COCHCH ₃₀; 1 and 0.9 (6H, 2d, CH(CH ₃)₂).

Intermediate 5: 2-(S), 3(S)dihydroxy-3(S)-hydroxycarbonyl-5 methylhexanoic, 1,1-dimethyl ester acid

a) 4-methyl-2-oxo pentanoic phenylmethyl ester acid

Disperse 78.7 g (0.51 mole) of compound a) of intermediary 4 in 500 mlof dry CH₂Cl₂. Add 39.2 ml (0.51 mole) of DMF. Cool to −20° C. and add45 ml (0.51 mole) of oxalyl chloride. Stir for 2 hours at roomtemperature.

At 0° C., add the mixture of 50 ml of CH₂Cl₂, 44 ml (0.425 mole) ofbenzyl alcohol, 143.4 ml of triethylamine. Stir for 18 hours at roomtemperature. Wash with HCl N, then NaHCO₃ (saturated solution). Dry overNa₂SO₄. Filter, evaporate to dryness. Distill at 114° C. under 3 mmHg.Recover 70.85 g (63%).

RMN (CDCl₃); δ7.45 (5H, m, H (Ar)); 5.3 (2H, s, OCH ₂Ar); 2.7 (2H, d, CH₂CO); 2.15 (1H, m, CH—(CH₃)₂); 0.95 (6H, d, CH(CH ₃)₂).

b) 5-methyl-3(phenylmethoxycarbonyl)(E) hex-2-enoic, dimethyl 1-1 ethylester acid

In a flask, introduce 46 g (0.10 mole) of tert-butoxycarbonyl methyltriphenyl phosphonium bromide. Add 12.8 g (0.1 05 mole) of tBuOK andstir for 30 minutes at room temperature. Add 20 g (0.091 mole) ofcompound a) diluted with 60 ml of DMF. Stir for one night at 20° C.

Evaporate to dryness.

Mix in isopropyl ether, filter, evaporate.

Purify by flash chromatography on 600 g of silica (eluant heptane:AcOEt;95:5).

Recover 19.9 g (69%) of product E.

RMN (CDCl₃): δ7.4 (5H, s, HAr); 6.75 (1H, s, COCH═); 5.2 (2H, s, OCH₂Ar); 2.75 (2H, d, CH ₂—C═); 1.9 (1H, m, CH—(CH₃)₂); 1.55 (9H, s, C(CH₃)); 0.95 (6H, d, CH(CH ₃)₂).

c) 2(S), 3(S) dihydroxy-5 methyl-3(S) phenylmethoxycarbonyl hexanoic,dimethyl 1,1 ethyl ester acid

Introduce into a flask 11.5 g of βAD-mix, 0.78 g (8.1 mmoles) of methylsulfonamide, 83 ml of a 1/1 mixture of tBuOH and H₂O. Stir for 2 minutesat +20° C., then cool to 0° C.

Add 2.6 g (8.1 mmoles) of compound b). Stir for 4 hours at 0° C., then 2hours at room temperature.

Add another 3 g of βAd-mix and stir for 1 night at 20° C. At 0° C., add16.4 g of sodium sulfite, stir for 1 hour at room temperature.

Extract with CH₂Cl₂ Wash with water, then with KOH 2N. Dry, filter,evaporate to dryness. Recover 3.2 g of oil (100%).

RMN (CDCl₃); δ7.4 (5H, s, HAr); 5.25 (2H, s, OCH ₂Ar); 4.2 (1H, d, OH);3.55 (1H, s, OH); 3.4 (1H, d, —CH—OH); 1.75 (3H, m, CH ₂—CH(CH₃)₂); 1.55(9H, s, OC(CH ₃)₃); 1 and 0.85 (6H, 2d, CH—(CH ₃)₂).

d) 2(S)[(S) 4′dimethylethoxycarbonyl-S-hydroxymethyl] 2S hydroxy 4methyl pentanoic acid

Solubilize 2 g of compound c) (5.66 mmoles) in 20 ml of methanol undernitrogen. Add 200 mg of 10% Pd/c. Purge with hydrogen and stir for 3hours at 20° C. Filter the catalyst on celite, evapaorate to dryness.

Recover 1.1 g (95%) of yellow solid.

MP: 110° C.

RMN (CDCl₃): δ5.9 (1H, very broad s, OH); 4.25 (1H, s, CHOH); 1.9 (2H,d, CH ₂—CH); 1.8 (1H, m, CH—(CH₃)₂); 1.55 (9H, s, (CH ₃)₃C); 1 and 0.9(6H, 2d, (CH ₃)₂ CH).

Intermediary 6: 3 (S) hydroxy-3 (S) hydroxycarbonyl-2(S) methoxy5-methylhexanoic, 1,1-dimethylethyl ester acid

a) 3(S)-hydroxy-2(S) methoxy-5 methyl 3(S) phenylmethoxycarbonylhexanoic, dimethyl 1.1 ethyl ester acid

To a suspension of 85 mg (3.4 mmoles) of NaH in 10 ml of dry THF, add at0° C. 1 g (2.8 mmoles) of compound c) of intermediary 5. Stir for 30minutes at 20° C.

At 0° C., add 0.9 ml (14 mmoles) of CH₃I. Stir for one night at roomtemperature.

Add HCl N. Extract with CH₂Cl₂ Dry, evaporate. Purify by flashchromatography (eluant heptane:AcOEt; 95:5).

Recover 380 mg (38%) of pure product.

RMN (CDCl₃): δ7.4 (5H, m, HAr); 5.25 (2H, dd, OCH ₂Ar); 3.85 (1H, s,CHOCH₃); 3.4 (1H, s, OH); 3.25 (3H, s, OCH ₃); 1.8 (2H, d, CH ₂CH); 1.7(1H, m, CH₂—CH—CH₃)₂); 1.55 (9H, s, C(CH₃)₃); 1 and 0.85 (6H, 2d, CH(CH₃)₂).

b) 3(S) hydroxy hydroxycarbonyl-2(S) methoxy-5 methyl hexanoic, dimethyl1.1 ethyl ester acid

To a suspension of 40 mg of 10% Pd/c in 5 ml of methanol, add 380 ml (1mmole) of the preceding compound. Stir for 2 hours at 20° C. underhydrogen atmosphere. Filter on celite. Rinse with methanol. Evaporate todryness. Recover 270 mg (93%).

RMN (CDCl₃): δ3.9 (1H, s, CH(OCH₃)); 3.45 (3H, s, OCH₃); 1.75 (2H, m, CH₂CH(CH₃)₂); 1.6 (10 H, m, C(CH ₃)₃+CH ₂CH); 1 (6H, dd, (CH(CH ₃)₂).

Intermediary 7: 2(S)[-2(S)-but-3-enyl]-2(S) hydroxy 4-methyl pentanoicacid

To 930 mg (2.67 mmoles) of compound d) of intermediary 4 in 8 ml ofethanol, add 8 ml of soda N (8 mmoles) and heat for one night underreflux.

Cool at 20° C., wash with ethyl ether. Acidify with HCl N and extractwith dichloromethane. Dry over Na₂SO₄, filter, evaporate.

Purify the 810 mg of oil obtained by flash chromatography on 37 g ofsilica (eluant: CH₂Cl₂:MeOH:AcOH; 95:5:0.5).

Recover 410 mg of pure product (82%).

[α]₃₆₅=+24.1° at t=20° C. (c=1.25, MeOH).

RMN (DMSO): δ5.7 (1H, m, CH₂═CH—); 5.05 (1H, d, CH ₂═); 5 (1H, s, CH₂═); 2.35 (1H, m, CH₂═CH—CH—CH₃); 1.65 (1H, m, CH—(CH₃)₂); 1.5 (2H, d,CH ₂—CH(CH₃)₂); 0.85 and 0.8 (9H, 3d, CH ₃).

Intermediary 8: 2-oxo-5-phenylpentanoic (S) ethoxycarbonyl (S)phenylmethyl ester acid

Synthesized in the same manner as for intermediary 4.

a) 2-oxo-5 phenyl pentanoic, sodium salt acid

RMN (DMSO): 7.2 (5H, m, H(Ar)); 2.5 (4H, m, COCH ₂CH₂CH ₂); 1.55 (2H, m,COCH₂CH ₂).

IR: ν ketone: 1706 cm⁻¹ ν COONa: 1625 cm⁻¹

b) 2-oxo-5 phenyl pentanoic (S) ethoxycarbonyl (S) phenyl methyl esteracid

RMN (CDCl₃): δ7.4 (10H, m, H(Ar); 6 (1H, s, OCHAr); 4.25 (2H, m, OCH ₂);2.95 (2H, m, ArCH ₂); 2.8 (2H, t, COCH ₂); 2.05 (2H, m, COCH₂CH ₂); 1.25(3H, t, CH₂CH ₃).

Intermediary 9: 2(S) hydroxy-3(R)methyl 2(S)-(2-propyl) pent-4-enoicacid

a) 2-oxo-3-methyl butyric (S) ethoxycarbonyl (S) phenylmethyl ester acid

Synethesized in the same manner as intermediary 4c.

RMN (CDCl₃): δ7.4 and 7.5 (5H, 2m, H(Ar)); 6.05 (1H, s, OCHAr); 4.2 (2H,m, OCH ₂CH₃); 3.3 (1H, m, CH(CH ₃)₂); 1.25 (9H, m, OCH₂CH ₃ and CH(CH₃)₂).

b) 2(S)hydroxy-3(R) methyl-2(S)-(2-propyl) pent-4-enoic(S)ethoxycarbonyl(S) phenyl methyl ester acid

This product was synthesized in the same manner as compound 4d.

RMN (CDCl₃): δ7.45 (5H, m, H(Ar)); 6 (1H, s, OCHAr); 5.85 (1H, m,CH₂═CH—); 5.75 (2H, m, CH ₂═CH; 4.25 (2H, m, OCH ₂CH₃); 3.1 (1H, broads, OH); 2.6 (1H, m, CH₂═CH—CH—CH₃); 2.15 (1H, m, CH(CH₃)₂); 1.3 (6H, 2d,CH₂═CH—CH ₃ and OCH₂CH ₃); 0.9 (6H, 2d, CH(CH ₃)₂).

c) Intermediary 9: 2(S) hydroxy-3(R) methyl 2(S)-(2-propyl) pent-4-enoicacid

This product was prepared in the same manner as intermediary 7.

RMN (CDCl₃); δ5.85 (1H, m, CH₂═CH—); 5.2 (2H, m, CH ₂═CH—); 3 (1H, verybroad s, OH); 2.75 (1H, m, CH₂═CH—CH—CH₃); 2.15 (1H, sept., CH(CH₃)₂);1.15 (3H, d, ═CHCH ₃); 1.05 (6H, 2d, CH(CH ₃)₂).

Intermediary 10: 4-chlorophenylalanine N-(2-methylthio-1-ethyl) amide

Solubilize in 5 ml of dry CH₂Cl₂ 500 mg (1.7 mmoles) of Boc4-chlorophenylalanine. Add 160 μl (1.7 mmoles) of2-methylthioethylamine, 415 mg (2 mmoles) of DCC and 270 mg (2 mmoles)of HOBT. Stir for one night at room temperature. Filter the DCU. Washwith HCl 1N, then NaHCO₃. Dry. Evaporate.

Take up with 6 ml of dry CH₂Cl₂. Add 1.5 ml of CF₃COOH and stir for 3hours at room temperature. Evaporate to dryness. Take up with AcOEt.Extract the product with HCl N. Neutralize the aqueous phase with NaHCO₃and extract with CH₂Cl₂. Dry. Evaporate. Recover 300 mg (or 74%) of pureproduct.

MP: 70° C.

RMN (CDCl₃): δ7.6 (1H, broad s, CONH); 7.3 and 7.5 (4H, 2d, H(Ar); 3.6(1H, m, NH₂CH); 3.55 (2H, q, CONHCH ₂); 3.25 (1H, 2d, CH ₂Ar); 2.75 (1H,m, CH ₂Ar); 2.65 (2H, t, CH ₂SCH₃); 2.15 (3H, s, SCH ₃); 1.3 (2H, s, NH₂).

The intermediaries 11 to 17 were synthesized in the same manner.

Intermediary 11: 4-chlorophenylalanine N-(2-(4-morpholino)-1-ethyl)amide

RMN (CDCl₃): δ7.35 (1H, broad s, CONH); 7.3 and 7.2 (4H, 2d, H(Ar)); 3.7(4H, m, —(CH ₂)₂O); 3.6 (1H, m, H₂N—CH—CO); 3.35 (2H, q, CONHCH ₂—);3.15 (1H, dd, CH ₂Ar); 2.75 (1H, dd, CH ₂Ar); 2.4 (6H, m, —CH ₂—N—(CH₂)₂—); 1.7 (2H, broad s, NH ₂).

Intermediary 12: 4-iodophenylalanine N-(2-methylthio-1-ethyl) amide

RMN (CDCl₃): δ7.65 (2H, d, H(Ar)); 7.5 (1H, broad s, CONH); 6.95 (2H, d,H(Ar)); 3.6 (1H, m, H₂NCHCO); 3.45 (2H, m, CONHCH ₂); 3.15 (1H, 2d, CH₂Ar); 2.7 (1H, 2d, CH ₂Ar); 2.6 (2H, m, —CH ₂SCH₃); 2.1 (3H, s, SCH ₃);1.7 (2H, broad s, H₂N).

Intermediary 13: 3-4-dichlorophenylalanine N-(2-methylthio-1-ethyl)amide

RMN (CDCl₃): δ7.55 (1H, broad s, CONH); 7.3 (2H, m, H(Ar)); 7.05 (1H, m,H(Ar)); 3.6 (1H, m, H₂NCHCO); 3.5 (2H, m, CONHCH ₂); 3.2 (1H, dd, CH₂Ar); 2.75 (1H, dd, CH ₂Ar); 2.6 (2H, m, CH ₂S); 2.15 (3H, s, SCH ₃);1.3 (2H, very broad s, H ₂N).

Intermediary 14: 4-chlorophenylalanine N-(2-cyano-1-ethyl) amide

RMN (CDCl₃): δ7.8 (1H, broad s, CONH); 7.3 (2H, d, H(Ar)); 7.15 (2H, d,H(Ar)); 3.65 (1H, m, H₂NCHCO); 3.55 (2H, m, CONHCH ₂—); 3.25 (1H, dd, CH₂Ar); 2.75 (1H, dd, CH ₂Ar); 2.65 (2H, t, CH ₂CN); 1.45 (2H, very broads, NH ₂).

Intermediary 15: 3,4-dichlorophenylalanine N-[2-(2 hydroxyethyl)oxyethyl] amide

RMN (CDCl₃): δ7.5 (1H, broad s, CONH); 7.3 (2H, m, H(Ar)); 7.1 (1H, m,H(Ar)); 3.75 (2H, m, CH ₂OH; from 3.65 to 3.45 (9H, m, H₂N—CH—CONHCH₂—CH₂O—CH ₂); 3.2 (1H, 2d, CH ₂Ar); 2.7 (1H, 2d, CH ₂Ar).

Intermediary 16: 3,4-dichlorophenylalanine N-[2-(2-methoxyethoxy)1-ethyl]amide

RMN (CDCl₃): δ7.5 (1H, broad s, CONH); 7.35 (2H, m, H(Ar)); 7.1 (1H, m,H(Ar)); from 3.45 to 3.7 (9H, m, H₂NCHCONH—CH ₂—CH ₂—O—CH ₂CH ₂—OCH₃);3.4 (3H, s, OCH ₃); 3.15 (1H, dd, CH ₂Ar); 2.9 (1H, dd, CH ₂Ar); 2 (2H,very broad s, NH ₂).

Intermediary 17: L βcyclohexylalanine N-(2-phenyl-1-ethyl)amide

RMN (CDCl₃); δ from 7.2 to 7.4 (6H, m, CONHCH₂ and H(Ar)); 3.55 (2H, m,H₂NCHCONHCH ₂); 3.4 (1H, dd, CONHCH ₂—); 2.85 (2H, m, CH ₂Ar); from 0.9to 1.9 (16H, 3 m, H ₂NCH—CH ₂-cyclohex.

Intermediary 18: D,L 2,4-dichlorophenylalanine N-methylamide

The racemic amino acids were synthesized by the methods known in thestate of the art.

Solubilize 1 g (4.27 mmoles) of D,L 2,4-dichlorophenylalanine in 35 mlof MeOH. Add 3.25 ml (25.6 mmoles) of TMSCl and heat for one night underreflux.

Evapaorate to dryness. Take up with 20 ml of MeOH. Freeze at −20° C. Add15 ml of methylamine and stir for 3 hours at +20° C. Evaporate todryness. Take up with water. Neutralize with NaHCO₃ and extract withchloroform. Dry. Evaporate. Recover 900 mg (or 85%) of oil.

RMN (CDCl₃): δ7.4 (1H, s, H(Ar)); 7.2 (2H, s, H(Ar)); 7.15 (1H, m,CONH); 3.7 (1H, m, NH₂CHCO); 3.45 (1H, dd, CH ₂Ar); 2.9 (1H, dd, CH₂Ar); 2.85 (3H, d, NHCH ₃); 1.55 (2H, broad s, NH₂).

The intermediaries 19 to 21 were synthesized in the same manner:

Intermediary 19: D,L 2,6-dichlorophenylalanine N-methylamide

RMN (CDCl₃): δ7.3 (2H, d, H(Ar)); 7.15 (1H, t, H(Ar)); 3.75 (1H, m,NH₂CHCO); 3.65 (1H, dd, CH ₂Ar); 3.15 (1H, dd, CH ₂Ar); 2.85 (3H, d,NHCH ₃).

Intermediary 20: D,L 3-chlorophenylalanine N-methylamide

RMN (DMSO): δ8.1 (1H, q, CONHCH₃); 7.25 (3H, m, H(Ar)); 7.15 (1H, d,H(Ar)); 3.55 (1H, m, CHCONHCH₃); 2.95 (1H, dd, CH ₂Ar); 2.75 (1H, dd, CH₂Ar); 2.55 (3H, d, NHCH₃).

Intermediary 21: D,L 2,5 dichlorophenylalanine N-methylamide

RMN (CDCl₃); δ from 7.15 to 7.35 (4H, m, H(Ar), CONH); 3.7 (1H, dd,H₂NCHCO); 3.5 (1H, dd, CH ₂Ar); 2.9 (4H, m, CH ₂Ar and NHCH ₃); 1.4 (2H,very broad s, NH₂).

Intermediary 22: 2 (R)[1(S*)(4-(methoxy)-benzylmercapto)ethyl]2(R*)-hydroxy 4-methylpentanoic acid

a) (E)-2[2(methyl)propyl]but-2-enoic ethyl ester acid

To 183.54 g (0.494 moles) of ethyltriphenylphosphonium bromide in 940 mlof THF, at room temperature, add 593 ml (0.593 mole) ofbis-trimethylsilyl sodium amide (1 M in THF), then 147 ml of HMPAdropwise.

Stir at room temperature for 45 minutes, then add 62.57 g (0.395 mole)of 4-methyl-2-oxopentanoic ethyl ester acid in solution in 60 ml of THFin 1 hour at room temperature. Stir for 1 hour at room temperature andpour the reaction medium over 900 ml of water and ice.

Extract with 3 times 800 ml of ethyl ether and dry over sodium sulfate,then evaporate under vacuum at 30° C.

Purify by flash chromatography (eluant: heptane:Et₂O; 99:1 then pentane:Et₂O; 97:3.

Recover 43.71 g of a yellow oil (65%).

RMN (CDCl₃): δ6.9 (q, 1H, CH═); 4.2 (q, 2H, OCH ₂CH₃); 2.2 (d, 2H, CH₂CH); 1.8 (d, 3H and m, 1H, CH ₃CH═ and CH₂CHCH₃); 1.3 (t, 3H, OCH₂CH₃); 0.9 (d, 6H, CH(CH ₃)₂);

IR (CHCl₃); νCO: 1701 cm⁻¹; νC═C: 1644 cm⁻¹.

b) 2 (S)[1 (oxoethyl)] 2-hydroxy 4-methylpentanoic ethyl ester acid

To 13.82 g (81.2 mmoles) of product a) in 1.12 l of acetone, 335 ml ofwater and 28.35 ml of acetic acid at −10° C., add with spatulas 22.32 g(141.2 mmoles) of KMnO₄.

Stir for 1 hour 30 minutes, then filter.

Evaporate the acetone from the filtrate, then extract with CH₂Cl₂ anddry over Na₂SO₄.

Evaporate under vacuum and purify by flash chromatography (eluant:heptane:CH₂Cl₂:AcOEt; 88:10:2).

Recover 3.53 g of a colorless liquid (21%).

RMN (CDCl₃): δ4.3 (q, 2H, OCH ₂CH₃); 4.2 (s, 1H, OH); 2.3 (s, 3H, CH₃CO); 2,1 (dd, 1H, CH ₂CH); 1.8 (m, 2H, CH ₂CH); 1.3 (t, 3H, OCH₂CH ₃);0.9 (dd, 6H, CH(CH ₃)₂).

IR (CHCl₃): νOH: 3510 cm⁻¹; νCO (ketone and ester): 1717 cm⁻¹.

c) 2 (S)[1(S*)-hydroxyethyl] 2(S*)-hydroxy 4-methylpentanoic ethyl esteracid

To 3.31 g (16.4 mmoles) of product b) in 33 ml of ethanol at 0° C., add0.62 g (16.4 mmoles) of sodium borohydride in 15 minutes. Stir for 5minutes, then evaporate the ethanol.

The residue is taken up in HCl 2N and extracted twice with Et₂O. Theethereal phases are dried over sodium sulfate, then evaporated. Purifyby flash chromatography (eluant: heptane: Et₂O: CH₂Cl₂; 70:20:10).

Recover 0.81 g of the least polar diastereoisomer (25%).

RMN (CDCl₃): δ4.3 (q, 2H, OCH ₂CH₃); 3.8 (m, 1H, CH—OH); 3.5 (s, 1H,OH); 2.3 (d, 1H, OH); 1.9 (dd, 1H, CH ₂—CH); 1.8-1.7 (m, 2H, CH ₂CH,CH(CH₃)₂); 1.4 (t, 3H, OCH₂CH ₃); 1.15 (d, 3H, CH—CH ₃); 1 (d, 3H, CH(CH₃)₂); 0.85 (d, 3H, CH(CH ₃)₂).

IR (CHCl₃): νOH: 3528 cm⁻¹; νCO: 1721 cm⁻¹.

d) 2 (S)[1-(S*)-ethylmethanesulfonate] 2-(S*)-hydroxy 4-methylpentanoicethyl ester acid

To 0.48 g (2.35 mmoles) of product c) dissolved in 10 ml of ethyl etherat 0° C., add 0.36 ml (2.58 mmoles) of triethylamine, then 0.2 ml (2.58mmoles) of methanesulfonic acid chloride. Stir for 1 night at roomtemperature.

The medium next is washed with H₂O, then soda 0.5 N, then water.

Dry over sodium sulfate and evaporate.

Recover 0.57 g of a viscous oil (86%).

RMN (CDCl₃): δ4.8 (q, 1H, CH—OSO₂CH₃); 4.3 (m, 2H, OCH ₂CH₃); 3.6 (s,1H, OH); 2.2 (dd, 1H, CH ₂CH); 1.9 (m, CH(CH₃)₂); 1.7 (dd, 1H, CH ₂CH);1.5 (d, 3H, CH—CH ₃); 1.4 (m, 3H, OCH₂CH ₃); 1 (dd, 6H, CH(CH ₃)₂).

IR (CHCl₃): νOH: 3518 cm⁻¹; νCO: 1728 cm⁻¹.

e) 2 (R)[1(S*)-(4-(methoxy)-benzylmercapto)ethyl]-2-(R*)-hydroxy4-methylpentanoic ethyl ester acid

To 0.33 g (2.12 mmoles) of 4-(methoxy)-benzylsulfide in 1.3 ml ethanol,add 0.97 ml (2.12 mmoles) of sodium ethylate (2.2 M in ethanol). Stirfor 5 minutes and add this solution to 0,54 g (1.91 mmoles) of productd) in 8.8 ml of ethanol. Stir for 4 hours at 70° C. then 1 night at roomtemperature.

Evaporate the ethanol and take up the residue in AcOEt. Wash with H₂O,NaOH 1N, then water.

Dry over sodium sulfate and evaporate.

Purify by flash chromatography (eluant: heptane:Et₂O; 98:2 then 97:3).

Recover 0.23 g (35%).

RMN (CDCl₃): δ7.2 (d, 2H, CHAr); 6.8 (d, 2H, CHAr): 4.3 (q, 2H, OCH₂CH₃); 3.8 (s, 3H, OCH ₃ and dd, 2H, CH ₂S); 3.4 (s, 1H, OH); 2.8 (q,1H, CHS); 2 (dd, 1H, CH ₂CH); 1.7-1.5 (m, 2H, CHCH ₂); 1.3 (t, 3H,OCH₂CH ₃); 1.25 (d, 3H, CH ₃CH); 1 (d, 3H, CH(CH ₃)₂), 0.9 (d, 3H, CH(CH₃ ₂).

IR (CHCl₃): νOH: 3526 cm⁻¹; νCO: 1723 cm⁻¹.

f) 2 (R) [1(S*) (4-methoxy) benzylmercapto)ethyl] 2 (R*) -hydroxy4-methylpentanoic acid

To 0.31 g (0.91 mmole) of product d) in 6 ml of ethanol, add 3.18 ml(3.18 mmoles) of soda 1N. Stir for 1 night at 80° C. Dilute with H₂O andextract with AcOEt. Acidify the aqueous phase with HCl 1N and extractwith CH₂Cl₂. Dry over sodium sulfate and evaporate. Crystallize byadding petroleum ether and a few drops of ethyl ether to the residue.Recover after filtration 0.123 g (43%).

RMN (CDCl₃): δ7.3 (d, 2H, CHAr); 6.8 (d, 2H, CHAr); 3.7 (s, 3H, OCH₃ anddd, 2H, CH ₂S); 3,3 (s, 1H, OH); 2.9 (q, 1H, CHCH₃); 2 (dd, 1H, CH ₂CH);1.8 (m, 1H, CH₂CH); 1.7 (dd, 1H, CH ₂CH); 1.3 (d, 3H, CH ₃CH); 1 (d, 3H,CH(CH ₃)₂), 0.9 (d, 3H, CH(CH ₃)₂).

Intermediary 23: 2(S) [1(S*)(o-benzylhydroxylamino)ethyl]2(S)-hydroxy4-methylpentanoic acid

a) 2 (S)[(1-(R)-hydroxyethyl)] 2-(S)-hydroxy 4-methyl pentanoic ethylester acid

To 21.84 g of βAD-mix in 70 ml of tert-butanol and 70 ml of waterstirred for 20 minutes, add 1.16 g (12 mmoles) of methane sulfonamide.

Cool to 0° C., then add 2.08 g (12 mmoles) of product a) of intermediary22.

Stir for 2 days at room temperature. Add 18 g of sodium sulfite and stirfor 2 hours.

Extract with AcOEt and wash the organic phase twice with KOH 2N.

Dry over sodium sulfate and evaporate.

Purify by flash chromatography (eluant: CH₂Cl₂:AcOEt:MeOH; 95:4:1).

Recover 1.88 g (77%).

RMN (CDCl₃): δ4.3 (q, 2H, OCH ₂CH₃); 3.9 (m, 1H, CHOH; 3.4 (s, 1H, OH);2.05 (d, 1H, OH); 1.7-1.5 (m, 3H, CH ₂CH); 1.3 (t, 3H, OCH₂CH ₃); 1.25(d, 3H, CH ₃CH); 1 (d, 3H, CH(CH ₃)₂); 0.9 (d, 3H, CH(CH ₃)₂).

IR (CHCl₃): νOH: 3518 cm⁻¹; νCO: 1724 cm⁻¹.

b) 2 (S)[1(oxoethyl)]2(S)-hydroxy-4-methylpentanoic ethyl ester acid

To 6.76 g (33.1 mmoles) of product a) in 200 ml of CH₂Cl₂ at 0° C., add9.4 ml (132 mmoles) of DMSO, then 18.7 g (132 mmoles) of P₂O₅.

Stir for 30 minutes at room temperature, then add another 9.38 g ofP₂O₅.

Stir for 16 hours at room temperature, then add 32.3 ml (231.7 mmoles)of triethylamine in 15 minutes.

Stir for 1 hour at room temperature, then add 200 ml of HCl 1N at 0° C.

Decant, wash the organic phase twice with HCl 1N. Purify by flashchromatography (eluant: heptane:AcOET; 95:5).

Recover 3.25 g of oil (48%).

RMN (CDCl₃): δ4.3 (q, 2H, OCH ₂CH₃); 4.2 (s, 1H, OH); 2.3 (s, 3H,CH₃CO); 2.1 (dd, 1H, CH ₂CH); 1.9 (dd, 1H, CH ₂CH); 1.8 (m, 1H, CH₂CH);1.3 (t, 3H, OCH₂CH ₃); 0.95 (dd, 6H, CH(CH ₃)₂).

IR (CHCl₃): νOH: 3504 cm⁻¹; νCO (ester+ketone): 1716 cm⁻¹.

c) 2 (S) [1(o-benzylhydroxylimino)ethyl] 2-(S)-hydroxy 4-methylpentanoicethyl ester acid

To 0.674 g (3.33 mmoles) of product b) in 10 ml of ethanol, add 0.585 g(3.66 mmoles) of o-benzylhydroxylamine hydrochlorate and 0.27 ml (3.33mmoles) of pyridine.

Stir at 95° C. for 3 hours.

Evaporate the ethanol and take up the residue with H₂O.

Extract with 3×50 ml of AcOEt.

Dry the organic phase and evaporate under vacuum.

Recover 1 g of a colorless oil (98%).

RMN (CDCl₃): δ7.3 (m, 5H, CHAr); 5.15 (s, 2H, CH ₂Ar); 4.2 (q, 2H, OCH₂CH₃); 3.9 (s, 1H, OH); 1.9 (m, 5H, CH ₃C═N and CH ₂CH); 1.8 (m, 1H,CH₂CH); 1.3 (t, 3H, OCH₂CH ₃); 0.95 (dd, 6H, CH(CH ₃)₂).

IR (CHCl₃): νOH: 3514 cm⁻¹; νCO: 1726 cm⁻¹.

d) 2 (S) [1(o-benzylhydroxyamino) ethyl] 2 (S)-hydroxy 4-methylpentanoicethyl ester acid

To 0.737 g (2.4 mmoles) of product c) in 20 ml of methanol, add 1.35 g(21.6 mmoles) of sodium cyanohydroboride, then 5.13 ml of hydrochloricmethanol dropwise.

Stir for 1 night and evaporate under vacuum.

The residue is taken up with HCl 1N and extracted three times withAcOEt.

Wash the organic phase wtih NaOH 1N, then H₂O.

Purify by flash chromatography (eluant: heptane:AcOEt; 9:1).

Recover 0.625 g of a colorless oil (84%).

RMN (DMSO): δ7.3 (m, 5H, HAr); 6.2 (d, 0.4H, NH); 6 (d, 0.6H, NH); 4.5(dd, 2H, CH ₂Ar); 4 m, 2H, OCH ₂CH₃); 3.25 (m, 0.6H, CHNH); 3 (m, 0.4H,CHNH); 1.7-1.4 (m, 3H, CH ₂CH); 1.2 (t, 3H, OCH₂CH ₃); 1.05 (dd, 2H, CH₃CH); 0.9 (dd, 3H, CH(CH ₃)₂); 0.8 (dd, 3H, CH(CH ₃)₂).

IR (CHCl₃): νOH: 3514 cm⁻¹; νCO: 1723 cm⁻¹.

e) 2 (S) [1(o-benzylhydroxylamino) ethyl]2-(S)hydroxy 4-methylpentanoicacid

To 0.6 g (1.95 mmoles) of product d) in 10 ml of ethanol, add 3.9 ml ofsoda 1N.

Stir at 100° C. for 6 hours. Evaporate the ethanol and take up theresidue with H₂O. Neutralize to pH 6 with H₂SO₄ 1 M.

Evaporate to dryness and take up the residue with methanol. Filter,evaporate.

Recover 0.55 g of a white foam (100%).

RMN (DMSO): δ7.3 (m, 5H, HAr); 6.6 (d, 0.4H, NH); 6.4 (d, 0.6H, NH); 4.9(s, 0.4H, OH); 4.8 (s, 0.6H, OH); 4.55 (m, 2H, CH ₂Ar); 2.9 (m, 1H,CHNH); 1.8-1.4 (m, 3H, CH ₂CH); 1 (dd, 3H, CH ₃CH); 0.9-0.8 (m, 6H,CH(CH ₃)₂).

IR (CHCl₃): νOH: 3416 cm⁻¹; νCO: 1724 cm⁻¹.

EXAMPLE 1 N-2(S)[1-hydroxycarbamoyl)-1-ethyl]2-(S) hydroxy-4methylpentanoyl O-methyltyrosine N methylamide A) N(1)[-2(S*)hydroxy-2(S*)(3 methyl) propyl 3(R*) methyl pent-4-enoyl]O-methyltyrosine N-methylamide

Solubilize 4.83 g (25.9 mmoles) of intermediary 1 in 100 ml of dryCH₂Cl₂. Add 5.40 g (25.9 mmoles) of o-methyltyrosine N-methylamide, 13.5g (25.9 mmoles) of PyBop and 11.3 ml (64.8 mmoles) ofdiisopropylethylamine and stir for 3 hours at room temperature. Washwith HCl N then NaHCO₃. Dry, evaporate.

Purify by flash chromatography on 800 g of silica (eluant:heptane:AcOEt; 50:50).

Recover 8.31 g (or 85%) mixture of diastereoisomers.

RMN (CDCl₃): δ7.3 (1H, m, CONH—); 7.15 (2H, d, CH(Ar)); 6.85 (2H, d,CHAr); 6.1 (1H, d, —CONH); 5.75 (1H, m, CH═); 5.1 (2H, m, CH ₂═); 4.6(1H, m, CH-α(Tyr)); 3.8 (3H, s, OCH ₃); 3.05 (2H, m, CH ₂Ar); 2.75 (3H,2d, CONHCH ₃); 2.5 (1H, m, C═C—CH—CH₃); 1.6 (3H, m, CH ₂—CH(CH₃)₂); 0.8(9H, m, CH ₃).

b) N-2(S*)[1(hydroxycarbonyl)-1 ethyl]-2(S*) hydroxy-4-methyl pentanoylO-methyltyrosine N-methylamide

Solubilize 8.05 g (21.4 mmoles) of product a) in 200 ml of tBuOH.Solubilize 45.7 g (214 mmoles) of NalO₄ in 845 ml of water. Add 0.68 g(4.3 mmoles) of KMnO₄, 3.25 g (23.5 mmoles) of K₂CO₃. Stir for ½ hour,then add 200 ml of tBuOH, then add the solution of product a) and stirfor 4 hours at room temperature.

Add HCl N and extract with 3 times 500 ml of AcOEt. Collect the organicphases and wash them with a thiosulfate acid solution. Dry over Na₂SO₄,evaporate to dryness.

Take up with NaOHN. Wash with ethyl ether. Acidify with HCl 6N andextract with AcOEt. Dry, evaporate. Recover 5,5 g (or 65%) (mixture ofdiastereoisomers).

RMN (DMSO): δ7.9 (1H, m, CONH); 7.6 (1H, m, CONH); 7.1 (2H, m, CH(Ar));6.8 (2H, m, CH(Ar)); 4.95 (1H, d, OH); 4.45 (1H, m, CH α(Tyr)); 3.7 (3H,s, OCH ₃); 2.9 (2H, m, CH ₂—Ar); 2.55 (4H, m, CONHCH ₃ and CHCOOH); from1.7 to 1 (3H, m, CH ₂—CH—(CH₃)₂); from 1 to 0.5 (9H, 3CH ₃).

c) N[2(S)[1(-phenylmethoxycarbamoyl)-1 ethyl]-2(S)hydroxy-4methylpentanoyl]O-methyl-L-tyrosine N-methylamide

Place 5.5 g (13.9 mmoles) of compound b) in 200 ml of dry THF. Add 3.16g (15.3 mmoles) of DCC, 2.07 g (15.3 mmoles) of HOBT, 4.45 g (27.9mmoles) of O benzylhydroxylamine hydrochlorate, 3.91 ml (27.9 mmoles) ofEt₃N and stir for one night at room temperature.

Filter the DCU and evaporate to dryness. Take up with CH₂Cl₂. Wash withHCl N, then saturated NaHCO₃. Dry, evaporate. Purify by flashchromatography on 300 g of silica (inject with: CH₂Cl₂; elute with:CH₂Cl₂:MeOH:NH4; 95:5:0.5).

Recover 3.34 g of the least polar pure diastereoisomer (50%).

RMN (CDCl₃):

the least polar diastereoisomer:

δ9.1 (1H, s, CONHO); 7.5 (1H, d, CONH); 7.4 (5H, s, Ar—CH₂—O); 7.2 (2H,d, CH(Ar)); 6.8 (2H, d, CH(Ar)); 5.9 (1H, q, CONHCH₃); 4.9 (2H, s,ArCHO); 4.8 (1H, s, OH); 4.4 (1H, m, CHH-α(Tyr)); 3.8 (3H, s, OCH₃);3.2-2.95 (2H, m, CH ₂Ar); 2.7 (3H, d, CONHCH ₃); 2.3 (1H, q,ONHCOCHCH₃); from 1.2 to 1.4 (3H, m, CH ₂CH(CH₃)₂); 0.95-0.8-0.7 (9H,3d, 3CH ₃).

the most polar diastereoisomer:

N(2R)[1(R) phenylmethoxycarbamoyl)ethyl] 2(R) hydroxy 4-methylL-tyrosine N-methylamide

RMN (CDCl₃): δ10 (1H, broad s, CONHOH); 7.7 (1H, d, CONH); 7.35 (5H, m,ArCH₂O); 7.1 (2H, d, H(Ar)); 6.8 (2H, d, H(Ar)); 6.2 (1H, broad q,CONHCH₃); 4.9 (3H, s+m, Ar—CH ₂—O+OH); 4.45 (1H, m, CH-α(Tyr)); 3.8 (3H,s, OCH ₃); 3 (2H, m, CH₂Ar); 2.8 (3H, d, NHCH ₃); 2.5 (1H, q, CHCH₃);1.6 and 1.3 (3H, 2 m, CH ₂CH); 1-0.85 and 0.7 (9H, 3d, 3CH ₃).

d) N-2-(S)[1(S) hydroxycarbamoyl)-1 ethyl]-2(S) hydroxy-4-methylpentanoyl O methyl L-tyrosine N-methylamide

3.28 g (6.8 mmoles) of the least polar diastereoisomer c) aresolubilized in 40 ml of absolute ethanol under nitrogen. Add 300 mg of10% Pd/c. Place under hydrogen atmosphere and stir for 2 hours at roomtemperature.

Filter the catalyst, rinse it with ethanol, evaporate to dryness.

Purify by flash chromatography on 200 g of silica (eluant: CH₂Cl₂:MeOH;95:5).

Recover 1.81 g (or 65%) of product.

[α]_(D)=+20.9° C. at t=20° C. (c=1, MEOH).

RMN (CDCl₃): δ10.75 (1H, s, HONHCO); 9.05 (1H, s, HONHCO); 7.9 (1H, q,CONHCH₃); 7.55 (1H, d, CONH); 7.05 (2H, d, CH(Ar)); 6.75 (2H, d,CH(Ar)); 5.42 (1H, s, OH); 4.5 (1H, m, CHα(Tyr)); 3.7 (3H, s, OCH ₃);2.6 (3H, d, CONHCH ₃); 2.3 (1H, q, HONHCOCH—CH₃); 1.55 (2H, m, CH₂CH(CH₃)₂); 1.22 (1H, m, CH ₂—CH(CH₃)₂); 0.85-0.7-0.6 (9H, 3d, 3CH₃).

EXAMKPLE 2 N-2-(S)[1(S)(hydroxycarbamoyl) ethyl]-2(S)hydroxy-4-methylpentanoyl 3-methyl-(S) valine N-methylamide a)N-[2(S)[1(S)((1,1 dimethyl) ethoxycarbonyl)ethyl]-2 (S) hydroxy4-methylpentanoyl] 3 methyl (S) valine N-methylamide

Solubilize 500 mg (1.9 mmoles) of intermediary 4 in 25 ml of dry CH₂Cl₂.Add 277 mg (1.9 mmoles) of (S) methyl 3-valine N-methylamide, 1.05 g ofPyBop and 744 μl (4 mmoles) of diisopropylethylamine. Stir for one nightat room temperature. Wash with HCl N, then saturated NaHCO₃. Dry,evaporate. Take up with AcOEt.

Filter on a silica gel. Evaporate.

Recover 598 mg (80%) of product.

RMN (CDCl₃): δ7.6 (1H, q, CONHCH₃); 6.1 (1H, broad d, CONH); 4.2 (1H, s,OH); 4.15 (1H, d, NH—CH—tBu); 2.75 (4H, m, NHCH ₃ and tBuOCOCH); 1.85(1H, m, CH ₂CH); 1.7 (1H, m, CH₂CH(CH₃)₂); 1.5 (9H, s, (CH ₃)₃CO); 1.4(1H, m, CH ₂—CH); 1.15 (3H, d, COCHCH ₃); 1.1 (9H, s, CH—C(CH ₃)₃); 0.95and 0.8 (6H, 2d, CH—(CH ₃)₂).

b) N 2(S) [1(S) (hydroxycarbonyl)ethyl)-2(S) hydroxy-4 methylpentanoyl]3 methyl (S) valine N-methylamide

Solubilize 580 mg of compound a) in 6 ml of dry CH₂Cl₂. Add 6 ml oftrifluoroacetic acid. Stir for 5 hours at room temperature. Evaporate todryness.

Take up 3 times with 10 ml of CH₂Cl₂ and 10 ml of heptane and evaporateto dryness.

Recover 480 mg of pure product (100%).

RMN (DMSO): δ8.05 (1H, q, CONHCH₃); 7.05 (1H, d, CONH); 5.2 (1H, verybroad s, COOH); 4.1 (1H, d, NHCHCO); 2.7 (1H, q, HO₂CCHCH₃); 2.55 (3H,d, NHCH ₃); 1.6 (2H, m, CH ₂—CH); 1.45 (1H, m, CH ₂—CH); 1 (3H, d,CO—CH—CH ₃); 0.9 (9H, s, C—(CH ₃)₃); 0.8 and 0.65 (6H, 2d, CH—(CH ₃)₂).

c) N-2-(S) [1(S) (hydroxycarbamoyl ethyl]-2 (S) hydroxy-4-methylpentanoyl 3 methyl-(S) valine N-methylamide

Solubilize 430 mg (1.3 mmoles) of compound b) in 9 ml of DMF. Add 204 mg(1.74 mmoles) of O-THP hydroxylamine, 195 mig (1.45 mmoles) of HOBT, 160μl (1.45 mmoles) of N-methylmorpholine, 284 mg (1/48 mmoles) of WSC, HCland stir for one night at room temperature.

Evaporate to dryness.

Take up with 10 ml of THF and 10 ml of HCl N and stir for 1 hour at roomtemperature.

Extract with AcOEt. Dry, evaporate.

Crystallize in ethyl ether. Recover 205 mg (or 45%).

RMN (DMSO): δ10.8 (1H, broad s, HONHCO); 9.1 (1H, broad s, HONHCO); 8.05(1H, q, CONHCH₃); 7.45 (1H, d, CONH); 5,5 (1H, s, OH); 4.15 (1H, d,NHCHCO); 2.6 (3H, d, NHCH ₃); 2.5 (1H, q, COCHCH₃); 1.6 (2H, m, CH₂CH(CH₃)₂); 1.3 (1H, m, CH ₂CH(CH₃)₂); 1 (3H, d, CH ₃CHCO); 0.9 (9H, s,C(CH ₃)₃); 0.85 and 0.65 (6H, 2d, —CH—(CH ₃)₂).

EXAMPLE 3 N-2 (S) [(S) hydroxy (S) hydroxycarbamoyl methyl] 2(S) hydroxy4-methyl pentanoyl L-4-chlorophenylalanine N-methylamide a) N-2-(S) [(S)1,1 dimethylethoxycarbonyl (S) hydroxymethyl] 2(S) hydroxy 4-methylpentanoyl L-4 chlorophenylalanine N-methylamide

To 495 mg (1.9 mmoles) of intermediary 5 in 5 ml of dry dichloromethane,add a catalytic quantity of DMAP, 1 ml (5.8 mmoles) of DIPEA, then 0.74ml (5.8 mmoles) of TMSCl. Stir for 30 minutes at 20° C., then add 255 mg(1.9 mmoles) of HOBT, 1 g (2 mmoles) of PyBop, 400 mg (2 mmoles) of4-chlorophenylalanine N-methylamide and 1 ml (5.8 mmoles) of DIPEA. Stirfor one night at 20° C. under nitrogen.

Add 385 mg (2 mmoles) of citric acid solubilized in 35 ml of methanoland stir for 30 minutes at 20° C.

Evaporate to dryness. Take up with AcOEt. Wash with HCl N, then NaHCO₃.Dry, evaporate.

Purify by flash chromatography (eluant: heptane:AcOEt; 40:60).

Recover 500 mg (58%) of pure product. MP: 168° C.

RMN (CDCl₃): δ7.3 (2H, d, H(Ar)); 7.2 (2H, d, H(Ar)); 6.9 (1H, d, NHCO);6.4 (1H, q, CONHCH₃); 4.6 (1H, m, NHCHCO); 4.1 (1H, d, CHOH); 4(1H, s,OH); 3.65 (1H, d, OH; 3.15 (2H, m, CH ₂Ar); 2.75 (3H, d, NHCH ₃); 1.65(3H, m, CH ₂CH); 1.55 (9H, s, (CH ₃)₃C); 1 and 0.85 (6H, 2d, (CH ₃)₂CH).

b) N-2-(S)[(S)hydroxy (S) hydroxycarbonyl methyl] 2(S) hydroxy4-methylpentanoyl L-4-chlorophenylalanine N-methylamide

Solubilize 500 mg of compound a) in 5 ml of dry CH₂Cl₂. Add 1.25 ml oftrifluoroacetic acid and stir for one night at 20° C. Evaporate todryness.

Take up with a CH₂Cl₂ 50/heptane: 50 mixture and evaporate to dryness.Recover 440 mg or 100%.

RMN (CDCl₃): δ7.9 (1H, d, CONHCH); 7.3 (2H, d, CH(Ar)); 7.15 (2H, d,CH(Ar)); 5.65 (1H, q, CONHCH₃); 4.55 (1H, m, NHCHCO); 4 (1H, s, CHOH);3.1 (2H, m, CH ₂Ar); 2.8 (3H, d, NHCH ₃); 1.75 (1H, m, CH(CH₃)₂); 1.65(2H, m, CH ₂CH); 1 and 0.8 (6H, 2d, CH(CH ₃)₂);

c) N-2-(S)[(S) hydroxy (S) hydroxycarbamoyl methyl] 2(S) hydroxy4-methyl pentanoyl L-4 chlorophenylalanine N-methylamide

Solubilize 440 mg (1.1 mmoles) of compound b) in 8 ml of dry DMF. Add at0° C. 180 mg (1.5 mmoles) of hydroxylamine OTHP, 180 mg (1.3 mmoles) ofHOBT, 150 μl (1.3 mmoles) of N-methylmorpholine, 250 mg (1.3 mmoles) ofWSC. Stir for one night at 20° C. Evaporate to dryness. Take up with 4ml of THF and 4 ml of HCl N and stir for 4 hours at 20° C.

Concentrate to dryness. Purify by flash chromatography (eluant: CH₂Cl₂:MeOH:AcOEt; 90:10:1).

Recover 160 mg (35%) of pure product. MP: 185° C.

RMN (DMSO): δ10.7 (1H, broad s, HONHCO); 9 (1H, broad s, HONHCO); 8.15(1H, q, CONHCH₃); 7.65 (1H, d, CONH); 7.25 (2H, d, H(Ar)); 7.20 (2H, d,H(Ar)); 6 and 5 (2H, 2 very broad s, OH); 4.45 (1H, m, COCHNH); 3.85(1H, s, CHOH); 3.05 (1H, dd, CH ₂Ar), 2.95 (1H, dd, CH ₂Ar); 2.5 (3H, d,NHCH ₃); 1.55 (1H, dd, CH ₂—CH); 1.4 (1H, m, CH₂CH); 1.25 (1H, dd, CH₂CH); 0.8 and 0.6 (6H, 2d, CH(CH ₃)₂).

Examples 4 to 28 were synthesized in the same manner as example 2, withtthe exception of example 7, synthesized in the same manner as example 1.

EXAMPLE 4 N-2(S)[1(S)(hydroxycarbamoyl)ethyl] 2(S) hydroxy4-methylpentanoyl L-phenyl alanine N-methylamide

M.P.: 174.2° (dec)

RMN (DMSO): δ10.75 (1H, broad s, HONHCO); 9.05 (1H, broad s, HONHCO);7.95 (1H, q, CONHCH₃); 7.60 (1H, d, CONH); 7.20 (5H, m, H(Ar)); 5.40(1H, broad s, COH); 4.45 (1H, m, NHCHCO); 2.95 (2H, dd, CH ₂Ar); 2.60(3H, d, NHCH ₃); 2.30 (1H, q, COCHCH₃); 1.60 (1H, ddd, CH(CH₃)); 1.55(1H, dd, CH ₂CH); 1.20 (1H, m, CH ₂CH); 0.85 (3H, d, CH ₃CH); 0.70 and0.50 (6H, 2d, CH(CH ₃)₂).

EXAMPLE 5 N-2(S)[-1-(S)(hydroxycarbamoyl)-ethyl]-2(S) hydroxy-4-methyl2entanol O methyl-L-tyrosine phenylmethylamide

RMN (DMSO): δ10.75 (1H, broad s, HONHCO); 9.05 (1H, broad s, HONHCO);8.5 (1H, t, NHCH₂Ar); 7.65 (1H, d, CONH); 7.25 (2H, m, CH(Ar)); 7.1 (5H,m, CH(Ar)); 6.8 (2H, d, CH(Ar)); 5.4(1H, s, OH); 4.6 (1H, m, NHCHCO);4.25 (2H, m, NHCH ₂Ar); 3.7 (3H, s, OCH ₃); 2.9 (2H, m, CHCH ₂Ar); 2.3(1H, q, COCHCH₃); 1.55 (2H, m, CH ₂CH); 1.2 (1H, dddd, CH(CH₃)₂);0.85-0.7 and 0.55 (9H, 3d, CH ₃CH and CH(CH ₃)₂).

EXAMPLE 6 N-2(S)[-1(S) (hydroxycarbamoyl) 1(S) ethyl]-2(S) hydroxy4-methyl pentanoyl leucinyl glycine methyl ester

RMN (DMSO): δ10.8 (1H, broad s, HONHCO); 9.1 (1H, broad s, HONHCO); 8.5(1H, t, CONH); 7.55 (1H, d, CONH); 5.45 (1H, s, OH); 4.45 (1H, m, CHCO);3.85 (2H, d, NHCH ₂CO); 3.60 (3H, s, OCH ₃); 2.45 (1H, q, COCH—CH₃);from 1.8 to 1.35 (4H, m, CH ₂—CH(CH₃)₂ and CH ₂CH); from 1 to 0.6 (12H,m, —CH(CH₃)₂ twice); 0.6 (3H, d, COCHCH ₃).

EXAMPLE 7 N-2(S) [-1-(S) hydroxycarbamoyl 1(S) ethyl] 2(S) hydroxy-3methyl butanoyl O-methyltyrosine N-methylamide

Synthesizd with intermediary 9.

RMN (DMSO): δ10.83 (1H, broad s, OHNHCO); 9.05 (1H, broad s, HONHCO);7.9 (1H, q, NHCH₃); 7.4 (1H, d, NHCH); 7.1 (2H, d, H(Ar)); 6.8 (2H, d,H(Ar)); 5.60 (1H, s, OH); 4.48 (1H, m, NHCHCO); 3.7 (3H, s, OCH ₃); 2.85(2H, 2dd, CH ₂Ar); 2.6 (1H, q, CHCH₃); 2.58 (3H, d, NHCH ₃); 1.75 (1H,m, CH(CH₃)₂); 0.85 and 0.8 (6H, 2d, CH(CH ₃)₂); 0.65 (3H, d, CHCH ₃).

EXAMPLE 8 N-2(S)[1(S)(hydroxycarbamoyl)-1 ethyl] 2(S) hydroxy 4-methylpentanoyl L-4-nitrophenylalanine N-methylamide

Dec.: 207° C. (dec)

[α]_(D)=+17.7° at t=20° C. (c=1, MeOH).

RMN (DMSO): δ10.7 (1H, broad s, HONHCO); 9.1 (1H, broad s, HONHCO); 8.21(2H, d, H(Ar)); 7.95 (1H, q, CONHCH₃); 7.7 (1H, d, CONH); 7.5 (2H, d,H(Ar)); 5.4 (1H, s, OH); 4.65 (1H, dd, NHCHCO); 3.1 (2H, d, CH ₂Ar); 2.6(3H, d, NHCH ₃); 2.25 (1H, q, COCHCH₃); from 1.7 to 1.4 (2H, m, CH ₂CH);1.2 (1H, dd, CH ₂CH); 0.85 (3H, d, CHCH ₃); 0.75 and 0.45 (6H, 2d, CH(CH₃)₂).

EXAMPLE 9 N-2(S)[1(S) (hydroxycarbamoyl) 1(S) ethyl]2-(S)hydroxy-4-methyl pentanoyl L-4-amino phenylalanine N-methylamide

RMN (DMSO): δ7.82 (1H, q, CONHCH₃); 7.50 (1H, d, CONH); 6.80 (2H, d,H(Ar)); 6.40 (2H, d, H(Ar)); 4.85 (2H, broad s, NH₂); 4.40 (1H, dd,NHCHCO); 2.70 (2H, d, CH ₂Ar); 2.55 (3H, d, CH ₃NH); 2,2 (1H, q,COCHCH₃); from 1.7 to 1.45 (2H, m, CH ₂CH); 1.25 (1H, m, CH ₂CH); 0.85(3H, d, CHCH ₃); 0.70 (6H, d, CH(CH ₃)₂).

EXAMPLE 10 N-2(S) [-1(S) (hydroxycarbamoyl)-1 ethyl]-2(S)hydroxy-4-methyl pentanoyl L4-chlorophenylalanine N-methylamide

[α]_(D)=+17.8° at t=20° C. (c=1, MeOH)

RMN (DMSO): δ10.75 (1H, broad s, HONHCO); 9.05 (1H, HONHCO); 7.9 (1H, q,CONHCH₃); 7.6 (1H, d, CONH); 7.25 (2H, d, 2HAr); 7.2 (2H, d, 2H(Ar));5.4 (1H, broad s, OH); 4.5 (1H, dd, COCHNH); 2.9 (2H, d, CH₂Ar); 2.55(3H, d, NHCH ₃); 2.25 (1H, q, CHCH₃); 1.55 (1H, m, CH ₂CH); 1.5 (1H, m,CH₂CH); 1.2 (1H, m, CH₂CH); 0.85 (3H, d, CHCH ₃); 0.65 and 0.5 (6H, 2d,CH(CH ₃)₂).

EXAMPLE 11 N-2(S)[-1(S)(hydroxycarbamoyl)-1 ethyl]-2(S) hydroxy 4-methylpentanoyl L-4-bromophenylalanine N-methylamide

MP: 214° C. (dec.)

RMN (DMSO): δ10.75 (1H, broad s, OHNHCO); 9.05 (1H, broad s, HONHCO);7.95 (1H, q, CONHCH₃); 7.65 (1H, d, CONH); 7.4 (2H, d, H(Ar)); 7.1 (2H,d, 2H(Ar)); 5.4 (1H, s, OH); 4.5 (1H, dd, NHCHCO); 2.9 (2H, d, CH₂(Ar)); 2.55 (3H, d, NHCH₃); 2.25 (1H, q, CHCH ₃); 1.5 (2H, m, CH ₂CH);1.2 (1H, m, CH ₂CH); 0.8 (3H, d, CHCH ₃); 0.65 and 0.5 (6H, 2d, CH(CH₃)₂).

EXAMPLE 12a N-2(S)[-1(S) hydroxycarbamoyl-1(S) ethyl-2(S)hydroxy-4-methyl pentanoyl L-4-chlorophenylalanineN-(2-(4-morpholino)1-ethyl)-amide

MP: 179° C. (dec.)

RMN (DMSO): δ10.75 (1H, broad s, OHNHCO); 9.05 (1H, broad s, HONHCO);7.95 (1H, broad t, CONHCH₂—); 7.65 (1H, d, CONH); 7.25 (4H, dd, H(Ar));5,4 (1H, s, OH); 4.5 (1H, m, CHCO); 3.55 (4H, m, (CH₂)₂—O); 3.15 (2H, q,CONHCH ₂); 2.9 (2H, dd, CH ₂Ar); 2.35 (4H, m, N(CH₂)₂); 2.3 (3H, m,CONH—CH₂—CH ₂, CHCH₃); 1.55 (2H, m, CH ₂CH); 1.2 (1H, dd, CH ₂CH); 0.85(3H, d, CHCH ₃); 0.7 and 0.55 (6H, 2d, CH(CH ₃)₂).

EXAMPLE 12b Hydrochlorate of example 12a

RMN (DMSO): δ10.8 (s, 1H, HONH); 10.7 (broad s, 1H, (CH₂)₃N⁺—H, Cl⁻);9.05 (s, 1H, HONH); 8.5 (m, 1H, CONHCH₂); 7.65 (d, 1H, CONH); 7.3 (m,4H, H(Ar)); 5.4 (s, 1H, OH); 4.55 (1H, m, NHCHCO); 4-3.6 (m, 4H, (CH₂)₂—O); 3.5-2.9 (m, 10H, CONHCH ₂ and (CH ₂)₃N⁺H, Cl⁻ and CH ₂Ar); 2.25(q, 1H, CHCH₃); 1.6 (m, 2H, CH ₂CH); 1.2 (m, 1H, CH₂CH); 0.85 (d, 3H,CHCH ₃); 0.7 and 0.5 (2d, 6H, CH(CH ₃)₂).

EXAMPLE 13 N-2(S)[-1-(S)(hydroxycarbamoyl)-1 ethyl]-2(S)hydroxy-4-methylpentanoylL-4-chlorophenylalanine N-(2methylthio-1-ethyl) amide

RMN (DMSO): δ10.7 (1H, broad s, HONHCO); 9.05 (1H, s, OHNHCO); 8.2 (1H,t, CONH); 7.6 (1H, d, CONH); 7.25 (4H, q, H(Ar)); 5.35 (1H, s, OH); 4.6(1H, m, NHCHCO); 3.2 (2H, dd, CH ₂—Ar); 2.9 (2H, m, NHCH ₂); 2.45 (2H,m, CH ₂S); 2.25 (1H, q, CHCH₃); 2.05 (3H, s, SCH ₃); 1.55 (2H, m, CH₂CH); 1.15 (1H, m, CH ₂CH); 0.85 (3H, d, CHCH ₃); 0.65 and 0.5 (6H, 2d,CH(CH ₃)₂).

EXAMPLE 14 N-2(S)[1(S) hydroxycarbamoyl-1 ethyl]-2(S) hydroxy 4 methylpentanoyl L-4-iodophenylalanine N-methylamide

RMN (DMSO): δ10.75 (1H, sl, OHNHCO); 9.05 (1H, broad s, HONHCO); 7.95(1H, q, CONHCH₃); 7.63 (1H, d, CONH); 7.6 (2H, d, H(Ar)); 7 (2H, d,HAr); 5.4 (1H, s, OH); 4.55 (1H, m, COCHNH); 2.88 (2H, dd, CH ₂Ar); 2.55(3H, d, NHCH ₃); 2.25 (1H, q, CHCH₃); 1.55 (2H, m, CH ₂CH); 1.2 (1H, m,CH ₂CH); 0.85 (3H, d, CHCH ₃); 0.7 and 0.5 (6H, 2d, CH(CH₃)₂).

EXAMPLE 15 N-2(S)[1(S) hydroxycarbamoyl-1 ethyl]2(S) hydroxy4-methylpentanoyl L-4-iodophenylalanine N-(2 methylthio-1-ethyl) amide

RMN (DMSO): δ10.7 (1H, broad s, OHNHCO); 9.1 (1H, broad s, HONHCO); 8.2(1H, dd, CONHCH₂); 7.6 (3H, d, H(Ar) and d, CONH); 7.05 (2H, d, H(Ar));5.3 (1H, s, OH): 4.6 (1H, m, NHCHCO); 3.3 (2H, m, CH ₂NH); 2.9 (2H, m,CH ₂Ar); 2.5 (3H, CH ₂S and CHCH₃); 2.1 (3H, s, SCH ₃); 1.6 (2H, m, CH ₂C); 1.2 (1H, m, CH ₂CH); 0.9 (3H, d, CH—CH ₃); 0.7 and 0.5 (6H, 2d,CH(CH₃)₂).

EXAMPLE 16 N-2(S)[1(S)hydroxy carbamoyl-1 ethyl]-2(S) hydroxy4-methylpentanoyl L-4-fluorophenylalanine N-methylamide

M.P.: 186.5° C.

RMN (DMSO): δ10.75 (1H, s, HONHCO); 9.05 (1H, broad s, HONHCO); 7.9 (1H,q, CONHCH₃); 7.65 (1H, d, CONHCH); 7.2 (2H, dd, H(Ar)); 7.05 (2H, dd,H(Ar)); 5.4 (1H, s, COH); 4.55 (1H, m, COCHNH); 2.9 (2H, d, CH ₂Ar); 2.6(3H, d, NHCH ₃); 2.3 (1H, q, CHCH₃); 1.6 (2H, m, CH ₂CH); 1.2 (1H, m, CH₂CH); 0.85 (3H, d, CHCH ₃); 0.7 and 0.55 (6H, 2d, CH(CH ₃)₂).

EXAMPLE 17 N-2(S) hydroxy 3(S) hydroxycarbamoyl 2(S) 1(2-methyl) propylhexanoyl L-4-chlorophenylalanine N-methyl amide

RMN (DMSO): δ7.9 (1H, q, CONHCH₃); 7.6 (1H, d, CONH); 7.2 (4H, m,H(Ar)); 5.3 (1H, broad s, OH); 4.5 (1H, m, NHCHCO); 2.9 (2H, d, CH ₂Ar);2.6 (3H, d, NHCH ₃); 2.15 (1H, q, CHCH₃); from 1.5 to 0.8 (7H, m, CH ₂CHand CHCH ₂CH ₂CH₃); 0.8 and 0.7 (9H, 1t and 2d, 3CH ₃).

EXAMPLE 18 N-2(S)[1(S)hydroxy carbamoyl-1 ethyl]-2 (S) hydroxy4-methylpentanoyl L-3,4-dichlorophenylalanine N-methylamide

MP: 216° C. (dec)

RMN (DMSO): δ10.7 (1H, s, HONHCO); 9.05 (1H, s, HONHCO); 7.95 (1H, q,CONHCH₃); 7.7 (1H, d, CONH); 7.5 (2H, 1d and 1s, H(Ar)); 7.2 (1H, d,H(Ar)); 5.45 (1H, s, OH); 4.55 (1H, m, NHCHCO); 2.9 (2H, d, CH ₂Ar); 2.6(3H, d, CONHCH ₃); 2.25 (1H, q, CHCH₃); 1.55 (2H, m, CH ₂CH); 1.2 (1H,m, CH ₂CH); 0.85 (3H, d, CHCH ₃); 0.6 and 0.5 (6H, 2d, CH(CH ₃)₂).

EXAMPLE 19 N-2(S)[1(S)hydroxy carbamoyl-1 ethyl]-2 (S) hydroxy4-methylpentanoyl L-3,4 dichlorophenylalanineN-hydroxyethyloxyethylamide

RMN (DMSO): δ8.1 (1H, t, NHCH₂); 7.7 (1H, d, CONH); 7.5 (2H, dd, H(Ar));7.25 (1H, dd, H(Ar)); 5.5 (1H, broad s, COH); 4.6 (1H, m, COCHNH); 3.5(2H, t, CH ₂OH, 3.4 (4H, m, CH ₂—O—CH ₂); 3.2 (2H, m, NHCH ₂); 3 (2H, d,CH ₂Ar)); 2.3 (1H, q, CHCH₃); 1.5 (2H, m, CH ₂CH); 1.2 (1H, m, CH ₂CH);0.85 (3H, d, CHCH ₃); 0.7 and 0.5 (6H, 2d, CH(CH ₃)₂).

EXAMPLE 20 N-2(S)[1(S)hydroxy carbamoyl-1 ethyl]-2 (S) hydroxy4-methylpentanoyl L-2,4-dichlorophenylalanine N-methylamide

The diastereoisomers were separated in step a (the least polardiastereoisomer)

MP: 193.5° C.

RMN (DMSO): δ10.8 (1H, s, CONHOH); 9.1 (1H, s, CONHOH); 7.9 (1H, q,CONHCH₃); 7.7 (1H, d, CONH); 7.5 (1H, s, H(Ar)); 7.4 (2H, 2d, H(Ar));5.4 (1H, s, OH); 4.6 (1H, m, COCHNH); 3.1 (2H, m, CH ₂(Ar)); 2.6 (3H, d,NHCH ₃); 2.3 (1H, q, CHCH ₃); 1.5 (2H, m, CH ₂CH); 1.2 (1H, m, CH₂CH);0.8 (3H, d, CHCH ₃); 0.7 and 0.5 (6H, 2d, CH(CH ₃)₂).

EXAMPLE 21 N-2(S)[1(S)hydroxycarbamoyl-1 ethyl]-2(S)hydroxy-4-methylpentanoyl L-3,4-dichlorophenylalanineN-(2-methylthio-1-ethyl) amide

RMN (DMSO): δ10.75 (1H, s, HONHCO); 9.05 (1H, s, HONH); 8.15 (1H, t,CONHCH₂); 7.7 (1H, d, CONHCH); 7.5 (1H, m, H(Ar)); 7.2 (1H, m, H(Ar));5.4 (1H, s, 0H); 4.6 (1H, m, NHCHCO); 3.25 (2H, q, CONHCH ₂); 2.95 (2H,m, CH ₂Ar); 2.4 (2H, m, CH ₂S); 2.28 (1H, q, COCHCH₃); 2.05 (3H, s, SCH₃); 1.55 (2H, m, CH ₂CH(CH₃)₂); 1.2 (1H, m, CH ₂CH(CH₃)₂); 0.85 (3H, d,COCHCH ₃); 0.65 and 0.5 (6H, 2d, CH(CH ₃)₂).

EXAMPLE 22 N-2(S)-[1(S) hydroxycarbamoyl-1 ethyl]-2(S)hydroxy-4-methylpentanoyl L-3-chlorophenylalanine N-methylamide

The diastereoisomers were separated in step a (the least polardiastereoisomer)

RMN (DMSO): δ10.75 (1H, s, HONHCO); 9.1 (1H, s, HONHCO); 7.95 (1H, q,CONHCH₃); 7.7 (1H, dd, CONHCH); from 7.3 to 7.1 (4H, m, H(Ar)); 5.4 (1H,s, COH); 4.55 (1H, q, NHCHCO); 2.9 (2H, d, CH ₂Ar); 2.6 (3H, d, NHCH ₃);2,24 (1H, q, COCHCH₃); from 1.7 to 1.45 (2H, m, CH ₂CH(CH₃)₂); 1.2 (1H,m, CH ₂CH(CH₃)₂); 0.85 (3H, d, COCHCH ₃); 0.7 from 0.5 (6H, 2d,CH(CH₃)₂).

EXAMPLE 23 N-2(S) [1(S) hydroxycarbamoyl-1 ethyl]-2(S) hydroxy 4-methylpentanoyl L-3,4-dichlorophenylalanineN-(2-(2-methoxyethoxy)1-ethyl)amide

RMN (DMSO): δ10.75 (1H, very broad s, OHNHCO); 9.05 (1H, broad s,HONHCO); 8.15 (1H, m, CONHCH₂); 7.72 (1H, d, CONHCH); 7.5 (1H, d,H(Ar)); 7.48 (1H, dd, H(Ar)); 7.25(1H, dd, H(Ar)); 5.45 (1H, s, COH);4.65 (1H, q, NHCHCO); from 3.5 to 3.2 (11H, m, CONHCH ₂CH ₂OCH ₂CH ₂OCH₃); 2.92 (2H, d, CH ₂Ar); 2.25 (1H, m, COCHCH₃); 1.55 (2H, m, CH₂CH(CH₃)₂); 1.2 (1H, m, CH ₂CH(CH₃)₂); 0.85 (3H, d, COCHCH ₃); 0.7 and0.5 (6H, 2d, CH(CH ₃)₂).

EXAMPLE 24 N-2(S)[1(S) hydroxycarbamoyl-1ethyl]-2(S)hydroxy-4-methylpentanoyl L-2,6-dichlorophenylalanineN-methylamide

The diastereoisomers were separated in step a (the least polardiastereoisomer)

RMN (DMSO): δ10.75 (1H, s, HONHCO); 9.1 (1H, s, HONHCO); 8.1 (1H, q,CONHCH₃); 7.65 (1H, d, CONHCH); 7.4 (2H, d, H(Ar)); 7.2 (1H, t, H(Ar));5.45 (1H, s, COH); 4.75 (1H, m, NHCHCO); 3.3 (1H, dd, CH ₂Ar); 3.1 (1H,dd, CH ₂Ar); 2.6 (3H, d, NHCH ₃); 2.2 (1H, q, COCHCH₃); 1.55 (2H, m, CH₂CH(CH₃)₂); 1.2 (1H, m, CH ₂CH(CH₃)₂); 0.85 (3H, d, COCHCH ₃); 0.6 and0.3 (6H, 2d, CH(CH ₃)₂).

EXAMPLE 25 N-2(S)[1(S) hydroxycarbamoyl-1ethyl]-2(S)hydroxy-4-methylpentanoyl L-2,5-dichlorophenylalanineN-methylamide

The diastereoisomers were separated in step a (the least polardiastereoisomer)

MP: 209-212° C.

RMN (DMSO): δ10.7 (1H, s, HONHCO); 9.1 (1H, s, HONHCO); 7.8 (1H, q,CONHCH₃); 7.7 (1H, d, CONHCH); 7.4 (2H, m, H(Ar)); 7.3 (1H, dd, H(Ar));5.4 (1H, s, COH); 4.6 (1H, m, NHCHCO); 3.0 (2H, m, CH ₂Ar); 2.6 (3H, d,NHCH ₃); 2.3 (1H, m, COCHCH₃); 1.5 (2H, m, CH ₂CH(CH₃)₂); 1.2 (1H, m, CH₂CH(CH₃)₂); 0.9 (3H, d, COCHCH ₃); 0.7 and 0.5 (6H, 2d, CH(CH ₃)₂).

EXAMPLE 26 N-2-(S)[1(S) hydroxycarbamoyl-1ethyl]-2(S)hydroxy-4-methylpentanoyl L-tryptophan N-methylamide

RMN (DMSO): δ10.75 (1H, s, HONHCO); 9.1 (1H, s, HONHCO); 7.95 (1H, q,NHCH₃); 7.65 (1H, d, CONHCH); 7.5 (2H, m, H(Ar)); 7.25 (1H, d, H(Ar));from 7.15 to 6.9 (3H, m, H(Ar)); 5.4 (1H, s, COH); 4.5 (1H, m, COCHNH);3 (2H, m, CH ₂Ar); 2.55 (3H, d, NHCH ₃); 2.2 (1H, q, COCHCH₃); from 1.65to 0.9 (3H, m, CH ₂CH(CH₃)₂); 0.75 (3H, d, COCHCH ₃); 0.6 and 0.4 (6H,2d, CH(CH ₃)₂).

EXAMPLE 27 N-2(S)[1(S) hydroxycarbamoyl-1ethyl]-2(S)hydroxy-4-methylpentanoyl L-4-chlorophenylalanineN-(2-cyano-1-ethyl) amide

IR: ν CN 2254 cm⁻¹

RMN (DMSO): δ10.75 (1H, s, HONHCO); 9.08 (1H, broad s, HONHCO); 8.5 (1H,q, CONHCH₃); 7.65 (1H, d, CONHCH); 7.25 (4H, dd, H(Ar)); 5.4 (1H, s,COH); 4.6 (1H, m, COCHNH); 3.2 (2H, m, CONHCH ₂CH₂); 2.95 (2H, m, CH₂Ar); 2.6 (2H, t, CH ₂CN); 2.25 (1H, q, COCHCH₃); 1.55 (1H, m, CH₂CH(CH₃)₂); 1.25 (2H, m, CH ₂CH(CH₃)₂); 0.95 (3H, d, COCHCH ₃); 0.65 and0.5 (6H, 2d, CH(CH ₃)₂).

EXAMPLE 28 N-2(S)[1(S) hydroxycarbamoyl-1ethyl]-2(S)hydroxy-4-methylpentanoyl L-4-chlorophenylalanineN-oximino-2-amino-2-ethylamide

This compound has been prepared from example 27.

Solubilize 130 mg (0.29 mmole) of compound of example 27 in 10 ml ofn-butanol, add 1.01 ml of a solution of hydroxylamine base 0.56 M (or0.57 mmoles) and heat for one night at 80° C. under nitrogen atmosphere;evaporate to dryness. Purify by flash chromatography on 10 g of silica(dry injection on 1 g of SiO₂). Elute CH₂Cl₂ 90:MeOH 10. Recover 74 mgor 53% of foam.

RMN (DMSO): δ10.7 (1H, broad s, HONHCO); 9 (1H, s, HONHCO); 8.8 (1H, s,C═NOH); 8 (1H, m, CONHCH₂); 7.62 (1H, m, CONHCH); 7.25 (2H, m, H(Ar));7.15 (2H, m, H(Ar)); 5.35 (2H, broad s, H ₂NC═NOH); 4.6 (1H, q, COCHNH);4 (1H, s, COH); 3.15 (2H, m, CONHCH ₂—); 2.85 (2H, m, CH ₂Ar); from 2.2to 2.02 (3H, m, COCHCH₃ and CH ₂—C═NOH); from 1.7 to 1 (3H, m, CH₂CH(CH₃)₂); 0.8 (3H, d, COCHCH ₃); 0.65 and 0.55 (6H, 2d, CH(CH ₃)₂).

EXAMPLE 29 N-2-(S)[(S) hydroxy-(S)hydroxycarbamoylmethyl)-2(S)hydroxy-4-methyl]pentanoyl O-methyl tyrosine N-methylamide

Synthesized in the same manner as example 3.

MP: 199° C. (dec.)

RMN (DMSO): δ10.72 (1H, s, HONH); 9 (1H, s, HONH); 8.2 (1H, q, CONHCH₃);7.6 (1H, d, CONH); 7.05 (2H, d, CH(Ar)); 6.8 (2H, d, CHAr); 6.1 (1H, d,OH); 4.85 (1H, s, OH); 4.38 (1H, m, NHCHCO); 3.9 (1H, d, CH—O); 3.7 (3H,s, OCH ₃); 2.92 (2H, 2dd, CH ₂Ar); 2.6 (3H, d, NHCH ₃); 1.55 (1H, dd, CH₂CH); 1.45 (1H, m, CH(CH₃)₂); 1.28 (1H, dd, CH ₂CH₂); 0.8 and 0.65 (2d,CH(CH ₃)₂).

EXAMPLE 30 N-2(S)[1(S)hydroxycarbamoylmethyl]-2(S)hydroxy-4-methylpentanoylL-3,4-dichlorophenylalanine N-methylamide

MP: 194.1° C.

RMN (DMSO): δ10.7 (1H, broad s, HONHCO); 9.0 (1H, s, HONHCO); 8.25 (1H,q, CONHCH₃); 7.7 (1H, d, CONHCH); 7.5 (1H, d, H(Ar)); 7.45 (1H, m,H(Ar)); 7.2 (1H, dd, H(Ar)); 6.05 (1H, d, COCHOH); 4.9 (1H, s, COH); 4.5(1H, m, COCHNH); 3.9 (1H, d, COCHOH); from 3.15 to 2.2 (2H, m, CH ₂Ar);2.6 (3H, d, CONHCH ₃); from 1.5 to 1.25 (3H, m, CH ₂CH(CH₃)₂); 0.8 and0.6 (6H, 2d, CH(CH ₃)₂).

EXAMPLE 31 N-2(S)[(S) hydroxycarbamoyl(S) methoxy]methyl 2(S) hydroxy-4methyl pentanoyl O-methyl tyrosine N-methylamide

Synthesized in the same manner as example 2 from intermediary 6.

RMN (DMSO): δ10.9 (1H, broad s, OHNHCO); 9.2 (1H, broad s, HONHCO); 8.05(1H, q, CONHCH₃); 7.55 (1H, d, CONH); 7.05 (2H, d, CH(Ar)); 6.8 (2H, d,CH(Ar)); 5.05 (1H, broad s, OH); 4.4 (1H, m, NHCHCO); 3.7 (3H, s, OCH₃);3.6 (1H, s, CHOCH₃); 3.2 (3H, s, CHOCH ₃); 2.9 (2H, m, CH ₂Ar); 2.6 (3H,d, NHCH ₃); 1.55 (3H, m, CH ₂CH); 0.8 and 0.65 (6H, 2d, CH(CH ₃)₂).

EXAMPLE 32 N-2-(S)[1(S)(hydroxycarbamoyl)ethyl]2(S) hydroxy 5-phenylpentanoyl L-3-cyclohexylalanine 2 phenylethylamide

Synthesized from intermediary 8.

RMN (DMSO): δ10.8 (1H, s, HONHCO); 9.1 (1H, s, HONHCO); 8.05 (1H, t,CONHCH₂); 7.5 (1H, d, CONH); from 7.3 to 7 (10H, m, H(Ar)); 5.45 (1H, s,OH); 4.35 (1H, m, NHCHCO); 3.18 (2H, q, NHCH ₂CH₂); 2.6 (2H, t, CH₂CH₂Ar); 2.5 (3H, m, CHCH₃ and CH ₂Ar); from 1.8 to 0.7 (17H, m: Hcyclohex, CH ₂-cyclohex; C(OH)CH ₂CH ₂); 1 (3H, d, CHCH ₃).

EXAMPLE 33 N-2-(S)[1(S)(hydroxycarbamoyl)ethyl]2(S) hydroxy5-phenylpentanoyl L-3,4-dichlorophenylalanine N-methylamide

Synthesized from intermediary 8.

RMN (DMSO): δ10.7 (1H, s, CONHOH); 9 (1H, s, CONHOH); 7.9 (1H, q,CONHCH₃); 7.7 (1H, d, CONH); 7.5 (2H, 1d and 1s, H(Ar)); from 7,2 to 7(6H, m, H(Ar)); 5.4 (1H, s, OH); 4.5 (1H, m, COCHNH); 2.9 (2H, m, CH₂Ar); 2.7 (1H, q, CHCH₃); from 2.5 to 2.3 (5H, m, CH₂CH ₂Ar and NHCH ₃);from 1.6 to 1.2 (4H, m, CH ₂CH ₂CH₂Ar); 0.5 (3H, d, CHCH ₃).

EXAMPLE 34N-2-(S)[(S)-hydroxy-(S)hydroxycarbamoylmethyl)-2(S)-hydroxy-4-methyl]pentanoyl-o-methyl-3-methyl-(S)valine N-methylamide

Synthesized in the same manner as example 3.

RMN (DMSO): δ9 (s, 1H, HONH); 8.05 (q, 1H, CONHCH₃); 7.5 (d, 1H, CONH);5.65 (d, 1H, OH); 5.35 (s, 1H, OH); 4.1 (d, 1H, NHCHCO); 3.9 (d, 1H,CHOH); 2.55 (d, 3H, NHCH ₃); 1.6 (m, 2H, CH ₂CH); 1.25 (m, 1H, CH ₂CH);0.9 and 0.7 (2d, 6H, CH(CH ₃)₂).

EXAMPLE 35a N-2(S)[1(S)-hydroxycarbamoyl-1 (S) ethyl]-2(S)-hydroxy-4-methyl pentanoyl L-(3,4)-dichlorophenylalanineN-(2-(4-morpholino)1-ethyl) amide

Synthesized in the same manner as example 2.

RMN (DMSO): δ10.7 (s, 1H, HONH); 9.05 (s, 1H, HONH); 8 (m, 1H, CONHCH₂);7.7 (d, 1H, CONH); 7.5 (m, 2H, H(Ar)); 7.2 (dd, 1H, H(Ar)); 5.4 (s, 1H,OH); 4.6 (m, 1H, NHCHCO); 3.5 (m, 4H, (CH₂)₂—O); 3.2 (m, 2H, CONHCH ₂);2.9 (d, 2H, CH ₂Ar); 2.4-2.2 (m, 7H, N(CH₂)₂ and CONHCH₂CH ₂ and CHCH₃);1.55 (m, 2H, CH ₂CH); 1.2 (m, 1H, CH₂CH); 0.8 (d, 3H, CHCH ₃); 0.7 and0.5 (2d, 6H, CH(CH ₃)₂).

EXAMPLE 35b Hydrochlorate of example 35a

RMN (DMSO): δ10.8 (m 1H, (CH₂)₃N⁺ H, Cl⁻); 10.7 (s, 1H, HONH); 9 (broads, 1H, HONH); 8.5 (m, 1H, CONHCH₂); 7.8 (d, 1H, CONH), 7.5 (m, 2H,H(Ar)); 7.2 (dd, 1H, H(Ar)); 5.4 (broad s, 1H, OH); 4.55 (m, 1H,NHCHCO); 4-3.4 (m, 6H, (CH ₂)₂—O and CONHCH); 3.1-2.9 (m, 8H, (CH₂)₃N⁺H, Cl⁻ and CH ₂Ar); 2.2 (q, 1H, CHCH₃); 1.5 (m, 2H, CH ₂CH); 1.2(m, 1H, CH ₂CH); 0.8 (3H, d, CHCH ₃); 0.65 and 0.45 (2d, 6H, CH(CH ₃)₂).

EXAMPLE 36 N-2(R)-[(1-thioethyl)] 2(R) hydroxy 4-methylpentanoyl-o-methyl-L-tyrosine methylamide a)2(R)-[1(4(methoxy)benzylmercapto)ethyl]2(R)hydroxy4-methylpentanoyl-o-methyl-L-tyrosine methylamide.

Synthesized in the same manner as compound a) of example 2 from theintermediary 22 and o-methyl-L-tyrosine methylamide.

RMN (CDCl₃): δ7.3-7.15 (m, 5H, HAr and CONH); 6.8 (m, 4H, HAr); 6 (m,1H, CONH); 4.7-4.5 (m, 1H, NHCHCO); 3.8 (2s and dd, 8H, 2OCH ₃ and CH₂S); 3.05 (m, 2H, CH ₂Ar); 2.9 (q, 1H, CHS); 2.7-2.6 (2d, 3H, CONHCH ₃);1.75 (d, 2H, CH ₂CH); 1.65 (m, 1H, CH₂CH); 1-0.75 (m, 9H, CHCH ₃ andCH(CH ₃)₂).

b) N-2(R)-[(1-thioethyl)] 2(R) hydroxy 4-methylpentanoyl-o-methyl-L-tyrosine methylamide

To 0.2 g (0.39 mmole) of compound a) in 6.3 ml of ammonia at −60° C.,add 56 mg (2.4 mmoles) of sodium. Stir for 10 minutes, then add ammoniumchloride. Allow the ammonia to evaporate by raising to room temperature.The residue is taken up in H₂O and extracted with CH₂Cl₂. The organicphase is dried on sodium sulfate and evaporated.

The diastereoisomers are separatred by flash chromatography (eluant:CH₂Cl₂:AcOEt; 80:20). Recover 28 mg of the least polar diastereoisomer(28%).

RMN (CDCl₃): δ7.3 (d, 1H, CONH); 7.2 (d, 2H, HAr); 6.8 (d, 2H, HAr);5.95 (m, 1H, CONHCH₃); 4.5 (m, 1H, NHCHCO); 3.8 (s, 3H, OCH ₃); 3.2 (m,1H, CHS); 3.05 (m, 2H, CH ₂Ar); 2.7 (d, 3H, NHCH ₃); 2.65 (s, 1H, OH);1.8-1.6 (m, 3H, CH ₂CH); 1.35 (d, 1H, SH); 1-0.8 (ddd, 9H, CH ₃CH andCH(CH ₃)₂).

IR (CHCl₃): νOH: 3619 cm⁻¹; νCO: 1675 cm⁻¹.

EXAMPLE 37 N-2 (R)-[1-thioethyl] 2(R) hydroxy 4-methylpentanoyl-L3,4)-dichlorophenylalanine a)2(R)-[1(4(methoxy)benzylmercapto)ethyl]2(R)-hydroxy4-methylpentanoyl-L-(3,4)-dichlorophenylalanine

Synthesized as example 34 a).

RMN (CDCl₃): δ7.3-6.9 (m, 8H, HAr and CONH); 6.3 (m, 0.7H, CONHCH₃); 6.1(m, 0.3H, CONHCH₃); 4.6 (m, 1H, NHCHCO); 3.9 (2s, 3H, OCH₃); 3.8 (m, 2H,CH ₂S); 3.2-2.8 (m, 2H, CH ₂Ar); 2.75 (2d, 3H, NHCH ₃); 1.7-1.5 (m, 3H,CH ₂CH); 1.3 (m, 3H, CH ₃CH); 1-0.7 (m, 6H, CH(CH ₃)₂).

b) N-2 (R)-[1-thioethyl] 2(R) hydroxy 4-methylpentanoyl-L-(3,4)-dichlorophenylalanine

Synthesized in the same manner as compound 35 b).

RMN (CDCl₃): δ7.4 (d, 1H, CONH); 7.3 (m, 3H, HAr); 6.1 (m, 1H, NHCH₃);4.6 (m, 1H, NHCHCO); 3.2-3 (m, 3H, CHS and CH ₂Ar); 2.7 (d, 3H, NHCH ₃);1.8-1.6 (m, 3H, CH ₂CH); 1.3 (d, 1H, SH; 1-0.8 (m, 9H, CH ₃CH, CH(CH₃)₂).

EXAMPLE 38N-2(S)-[1(N-hydroxy-N-formyl)ethyl]2(S)-hydroxy4-methylpentanoyl-o-methylL-tyrosine methylamide a) N-2(S)-[1(o-benzylhydroxylamino)ethyl]2(S)-hydroxy-4-methylpentanoyl-o-methyl-L-tyrosine methylamide

Synthesized in the same manner as compound a) of example 2 except forthe purification performed by flash chromatography (eluant:heptane:AcOEt; 20:80 then 30:70).

RMN (DMSO): δ7.9 (m 1H, CONH); 7.6 (d, 1H, CONH); 7.25 (m 5H, HAr); 7.05(dd, 2H, HAr); 6.75 (dd, 2H, HAr); 6.1 (d, 0.5H, NH); 5.95 (d, 0.5H,NH); 4.9 (s, 0.5H, OH); 4.85 (s, 0.5H, OH); 4.55 (s, 1H, OCH ₂Ar); 4.5(s, 1H, OCH ₂Ar); 4.45 (m, 1H, NHCHCO); 3.7 (s, 3H, OCH₃); 3.1 (m, 0.5H,CHNH); 3 (m, 0.5H, CHNH); 2.7 (m, 2H, CHCH ₂Ar); 2.55 (dd, 2H, NHCH ₃);1.7-1.3 (m, 3H, CH ₂CH); 0.95-0.6 (m, 9H, CHCH ₃ and CH(CH ₃)₂).

IR (CHCl₃): νOH: 3454 cm⁻¹; νNH: 3401 cm⁻¹; νCO: 1675 cm⁻¹.

b) N-2(S)-[1-(N-formyl (o-benzylhydroxylamino))ethyl] 2(S)-hydroxy-4-methylpentanoyl-o-methyl-L-tyrosine methylamide

To a solution of formic acid (85 μl; 2.24 mmoles) and acetic anhydride(0.22 ml; 2.24 mmoles) heated to 40° C. for 45 minutes, add 0.264 g(0.56 mmoles) of compound a) dissolved in 11 ml of CH₂Cl₂.

Stir at room temperature for 1 hour 30 minutes. Wash with an 80% NaHCO₃solution, then with H₂O. Dry over sodium sulfate and evaporate undervacuum.

Recover 0.267 g (95%).

The product is a ≈50-50 mixture of diastereoisomers. Eachdiastereoisomer exists in the form of two RMN conformers.

RMN (DMSO): δ8-7.6 (m, 3H, CHO and CONH and COCHCH₃); 7.4 (m, 5H, HAr);7 (m, 2H, HAr); 6.8-6.6 (m, 2H, HAr); 5.6-5.2 (m, 1H, OH); 5-4.5 (m, 3H,OCH ₂Ar and NHCHCO); 3.7-3.6 (2s, 3H, OCH ₃); 2.8 (m, 2H, CH ₂Ar); 2.6(m, 3H, NHCH ₃); 1.6 and 1.4 (m, 3H, CH ₂CH); 1.2 and 1 (d, 3H, CHCH ₃);0.8 and 0.65 (m, 6H, CH(CH ₃)₂).

IR (CHCl₃); νOH: 3454 cm⁻¹; νNH: 3393 cm⁻¹; νCO: 1661 cm⁻¹.

c) N-2(S)-[1(N-hydroxy-N-formyl)ethyl] 2(S)-hydroxy4-methylpentanoyl-o-methyl L-tyrosine methylamide

To a 10% suspension of Pd/C (0.266 g) in 5 ml of MeOH, add 0.266 g ofcompound b) (0.53 mmole) in solution in 25 ml of MeOH.

Stir under hydrogen for 1 hour. Filter on celite and evaporate undervacuum.

Purify by flash chromatography (eluant CH₂Cl₂:MeOH; 97:3).

Recover 0.118 g (54%).

The product is a ≈50-50 mixture of diastereoisomers. Eachdiastereoisomer exists in the form of two RMN conformers.

RMN (DMSO+D₂O): δ8.2 and 8.1 and 7.8 and 7.7 (4s, 1H, CHO); 7 (m, 2H,HAr); 6.7 (m, 2H, HAr); 4.5 (m, 1H, NHCHCO); 4.2 and 3.7 (m, 1H, CHCH₃);3.8 (s, 3H, OCH₃); 2.8 (m, 2H, CH ₂Ar); 2.6 (2d, 3H, NHCH ₃); 1.6-1.25(m, 3H, CH ₂CH); 1.2-0.6 (m, 9H, CHCH ₃ and CH(CH ₃)₂).

IR (CHCl₃): νOH: 3450 cm⁻¹; νNH: 3397⁻¹; νCO: 1664 cm⁻¹.

Biochemistry

Activity on Collagenase

The anti-collagenase activity of the compounds according to thisinvention is determined on the collagenase of the human monocytic stockU937 in accordance with the operating mode described by Cawston andBarrett (Anal. Biochem. 99, 340-345, 1979).

The collagenase was incubated at 27° C. for 16 hours with radiomarkedcollagen, and its activity was determined by the release of solublepeptides produced by the enzymatic breakdown of the collagen. Theinhibiting concentration C150 for each compound was determined bymeasurement of the activity of the enzyme in the presence of theinhibitor at a concentration ranging between 10⁻⁷ and 10⁻¹² M (theinhibitor is dissolved at a concentration of 10⁻² M in DMSO, thendiluted successively by tenths in the buffer: Tris, HCl 150 mM, pH 7.5;NaCl 0.15 M; CaCl₂ 10 mM).

Activity on Stromelysine

The anti-stromelysine activity of the compounds according to thisinvention was determined on stromelysine of human origin, in accordancewith the operating mode described by Cawston, T. E., Galloway, W. A.,Mercier, E., Murphy, G., Reynolds, J. J. (Biochem. J., 195, 159-165,1981). The stromelysine was incubated at 37° C. for 16 hours withradiomarked transferrin, and its activity was determined by the releaseof the soluble peptides produced by the enzymatic breakdown of thetransferrin. The inhibiting concentration C150 for each compound wasdetermined by measurement of the activity of the enzyme in the presenceof the inhibitor at a concentration ranging between 10⁻⁷ and 10⁻¹² M(the inhibitor is dissolved in the same manner as for collagenase).

Activity on Gelatinase

The anti-gelatinase activity of the compounds according to the inventionwas determined on 72 kDa gelatinase of human origin, in accordance withthe operating mode described by Knight, C. G., Willenbrock, F., Murphy,G. (FEBS Letters, 296 (3), 263-266, 1992).

The gelatinase was incubated at 37° C. for 10 minutes, and its activitywas determined by the increase in fluorescence of the product ofMca-Pro-Leu breakdown of the Mca-Pro-Leu-Gly-Leu-Dpa-Ala-Arg-NH₂synthetic substrate.

The inhibiting concentration C150 for each compound was determined bymeasurement of the activity of the enzyme in the presence of theinhibitor at a concentration ranging between 10⁻⁷ and 10⁻¹² M (theinhibitor is dissolved at a concentration of 10⁻² M in DMSO, thendiluted successively by tenths in DMSO to 10⁻⁴ M, then in the buffer:Tris, HCl 50 mM+0.05% Brij 35; CaCl₂ 10 mM; NaCl 0.15 M).

The anti-gelatinase activity of the compounds according to the inventionalso was determined on 92 kDa or 72 kDa gelatinase of human origin, inaccordance with the operating mode described by Harris, E. D., Krane, S.M.; (Biochem. Biophys. Acta, 258, 566-576, 1972).

The gelatinase was incubated at 37° C. for 16 hours with radiomarkedgelatin, and its activity was determined by the release of the solublepeptides produced by the enzymatic breakdown of the gelatin. Theinhibiting concentration C150 for each compound was determined bymeasurement of the activity of the enzyme in the presence of theinhibitor at a concentration ranging between 10⁻⁷ and 10⁻¹² M (theinhibitor is dissolved in the same manner as for collagenase).

Inhibition of the Production of TNF in vitro

The capacity of the compounds according to this invention to inhibit therelease of TNF was determined from the peritoneal macrophages of micestimulated by LPS (Lang, F., Robert, J. M., Boucrot, P., Welin, K,Petit, J. Y., J. Pharmacol. Exp. Ther., 275, 171-176, 1995).

After an adhesion phase of 2 hours, the cells were treated for 1 hourwith the compound to be tested at concentrations ranging between 10⁻⁶ to10⁻⁹ M (the inhibitor is dissolved in DMSO at 10⁻² M, then at 5.10⁻³ Min RPMI 1640 with 5% SVF, then diluted successively by tenths in thesame medium), then stimulated by the addition of LPS (100 ng/ml in finalconcentration). Three hours later, the biological activity of the TNF inthe supernatants was determined by a cytotoxicity test using the murinestock L929 in accordance with the operating procedure described by Band,G., Lin, C. W., Georgescu, H. I., Evans, C. H. (Bioch. Biophys. Acta,1134, 309-318, 1982).

The pg/ml concentration of the TNF of the samples was determined from amurine αTNF standard curve, thus making it possible to define theinhibiting concentration C150 of the compounds studied. The results oncollagenase, gelatinase, stromelysine and TNF are compiled in table 1.

Inhibition of the Breakdown of Cartilage in vivo

The activities of the compounds of example 1 and example 10comparatively to the compound BB16 were determined in a model ofbreakdown of cartilage in accordance with the operating mode describedby Bottomley, K. M. K., Griffith, S. R. J., Rising, T. G., Steward, A.(Brit. J. Pharmacol. 21, 287-289, 1988). Femoral head cartilage of therat, wrapped in sterile cotton, is implanted subcutaneously on the backsof mice. After 21 days, the mice are euthanized, the cartilagesrecovered and weighed. The animals are treated each day, twice a day,with one 10-mg/kg dose i.p.

The compound of example 10 inhibits weight loss of the cartilage by 65%,while the compound of example 1 inhibits the lattter in the same manneras comparator 1: BB16; that is, 42% (p<0.05).

Inhibition of Septic Shock

Septic shock is induced in female Balb/c mice in accordance with theprocedure described by Mohler, K. M. et al. (Nature, 370, 218-220,1994). Briefly, the shock is induced by i.p. injection of 20 mg ofD-galactosamine and i.v. injection of 20 ng of LPS. The survival of theanimals is evaluated for the following 48 hours.

The products to be tested are injected i.p. 30 minutes prior toinduction of the shock. The compounds according to the invention protectfrom shock from 10 to 50 mg/kg. For instance, the compound of example 10protects at 50 mg/kg.

Inhibition of Production of αTNF in vivo

The injection of LPS in mice (100 μg/mouse i.p.) induces the productionof αTNF, the plasmatic concentration of which is maximal at 60-90minutes (Sekut, L., Menius, J. A., Brackeen, M. F., Connolly, K. M., J.Lab. Clin. Med., 124, 6, 613-820, 1994).

αTNF is titrated by its cytotoxicity on L929 cells (Flick, D. A.,Gilford, G. E., J. Immunol. Methods, 68, 167-175, 1984).

The products to be tested are administered i.p. or p.o. prior toinjection of LPS.

The compounds according to the invention inhibit the production of αTNFbetween 1 and 50 mg/kg.

Inhibition of Release of αTGF

The compounds of this invention were tested in accordance with themethods of examples 2 and 3 described in patent WO 96/25156 and inhibitthe release of αTGF.

Determination of Bioavailablity

The compounds of this invention were administered i.p. or p.o. between 1and 50 mg/kg to mice: the blood levels then were determined by anex-vivo bioassay according to Wang X. et al. (Cancer Res., 54,4726-4728, 1994). The results are presented in Table 2. The introductionof an OH group systematically increases the bioavailability of theproducts.

TABLE 1 C150 (nM) gelatinase C150 collagen- 92 72 strome- (μM)Structure, JL ase kDa* kDa** lysine TNF

5 1 1 15 1

18 16 3.4 60 NT

50 54 6.7 >1000 3.2

36 10 1.6 29 NT

12 0.5 2.4 11 NT

24 21 8.5 45 NT

30 17 18 NT 2

7 1.4 1.9 28 ≦0.38

11 2 1.6 50 2.2

4 2 1 48 0.4

10 2 2 37 0.61

23 9 10 15 0.51

22 8 NT NT 1.6

10 0.005 1.6 20 0.15

2 0.2 0.3 12 0.4

32 0.001 1.3 38 4.5

14 2 1.2 NT 0.6

NT 1.6 1.6 NT 0.27

2 0.002 0.4 6 ≦0.14

7 0.001 0.7 NT 0.5

3.7 0.2 1.6 NT 1

5 83% NT NT 0.4

2.4 NT NT NT 0.13

NT 60% 0.14 NT 0.37

NT NT NT NT NT

NT NT NT NT NT

NT 0.6 NT NT 4.4

NT 1.7 NT NT NT

26 1.6 NT NT NT

26 80 13 30,000 NT

11 7 NT >1,000 3.6

120 140 18 90 NT

NT 1.9 0.8 NT >1

225 86% 0.01 NT 0.58

34 NT NT NT

6 0.38 NT NT 0.86

4 1.1 NT NT 0.3

500 23 NT NT 3

>1,000 70 NT NT 2

130 17 NT NT 3.2

5 7 0.6 10 0.14

4 11 0.5 47 0.14 *Test performed with radiomarked gelatin **: performedon synthetic substrate NT: not tested

TABLE 2 I.p. method (μg/ml) P.o. method (μg/ml) PRODUCT (50 mg/kg) 10′20′ 40′ 1 h 24 h 5′ 10′ 20′ 30′ 40′ 1 h 24 h

0.2 <0.1 <0.1 <0.1 <0.1 <0.1

23 8.7 0.77 <0.5 4.2 2.1 0.58 <0.5

30 27 18 2.3 1.2 0.9

87 13 1.7 0.3 5.1 3.1 0.4 <0.3

0.6 0.3 0.6

0.7 0.5 0.1 0.1 0.1

6 5 1.8 1.25 0.8 0.45 LD

4.5 7.8 4.4 0.1 0.1 LD

20 9 47 2.2 4.3 0.3 0.5 0.17

20 0 4 10 9 9 9

56 22 3.5 NT NT NT NT NT NT NT

What is claimed is:
 1. Compounds of the following general formula (X):

in which: Y represents: —CONHOH, or —SH, or a group with the formula

 or a group with the formula

in which: R₄ represents —H, or a C₁ to C₆ alkyl group, or a phenylalkylgroup in which the alkyl group is C₁ to C₆, R₅ represents a group withthe formula

in which: R₆ represents —H, or a C₁ to C₆ alkoxy group, or a benzyloxygroup, R₇ represents —H, or a halogen atom R₁ represents: a C₃ to C₁₆linear or branched, or C₃ to C₆ cyclized alkyl chain, said chaincomprising a heteroatom, a phenoxyalkyl or phenylalkyl group,substituted or unsubstituted, or a heteroarylalkyl group, the alkylgroup being C₂ to C₅, R₂ represents: a hydrogen atom, or, a C₁ to C₅alkyl or C₂ to C₅ alkylidene group, or a hydroxyl, a C₁ to C₆ alkoxy ora benzyloxy, provided that Y represents —CONHOH when R₂ represents ahydroxyl, or a hydroxymethyl, or C₁ to C₆ alkoxymethyl group, or anarylalkyl group in which the alkyl portion is C₁ to C₆, an aryloxymethylgroup, an arylthiomethyl group, a heteroarylthiomethyl group, in whicharyl designates a substituted or unsubstituted phenyl remainder —OCH₃, alinear or branched C₁ to C₃ alkyl group, a halogen, an amine group, or aphthalimide alkyl group in which the alkyl portion is C₁ to C₆, or analkoxycarbonmethyl group, a benzyloxycarbonylmethyl, an acetylmethyl,provided that Y represents —SH in these three cases, AA represents anamino acid, or an amino acid chain a group with the formula

R₃ represents a group with the formula —NH—(R₈)_(n)—R₉ in which: nrepresents 0 or 1, R₈ represents a linear or branched alkyl chain, with1 to 8 carbon atoms unsubstituted or substituted one or severalheteroatoms, R₉ represents a hydrogen atom or a methyl, nitrile,morpholino, phenyl, methoxy, hydroxyl, thiomethyl group, or a group withthe formula —CH(NH₂)═N—OH, or a —N(CH₃)₂ group.
 2. The compoundsaccording to claim 1, having the following formula (Xa):

in which: Y represents: —CONHOH, R₁ represents: —CH(CH₃)₂,—CH₂—CH(CH₃)₂, or

R₂ represents: an alkyl group with 1 to 5 carbon atoms, a hydroxyl, oran alkoxy group with 1 to 5 carbon atoms, R represents: —C(CH₃)₃,—CH₂—CH(CH₃)₂,

 aromatic or nonaromatic, in which R_(a) and R_(b), independently of oneanother, represent —H, —Cl, —Br, —I, —F, —OCH₃, —NO₂, —NH₂, or a groupwith the formula

R₃ represents a —NH—(CH₂)_(n1)—R₉ group in which: n₁ represents 0, 1 or2, R₉ represents —CH₃, —C≡N, —COOHCH₃, —SCH₃, —O—(CH₂)₂—OH,—O—(CH₂)₂—OCH₃, —CH(NH₂)═N—OH,


3. The compounds according to claim 2, wherein the R₁ and R₂substituents are positioned in anti orientation in relation to thesuccinic residue in accordance with the following formula (XI.1):


4. Compounds according to claim 1, wherein R represents a group with theformula

in which R_(a) and R_(b) represent a halogen atom.
 5. The compoundsaccording to claim 1, wherein R₃ represents a group with the formula—NH—(CH₂)₂—SCH₃.
 6. A mixture comprising compounds with the followingformula (XI.1):

in which Y, R₁ and R₂, AA and R₃ are such as defined in claim 20, andcompounds with the following formula (XI.2):

in which Y, R₁, R₂, AA and R₃ have the meaning indicated hereinabove,the proportion of the compounds (XI.1) and (XI.2) in the mixture beingapproximately 50% to approximately 99% for the compound of formula(XI.1) and approximately 50% to approximately 1% for the compound offormula (XI.2).
 7. The compounds according to claim 1, selected from thegroup consisting of:


8. A pharmaceutical composition comprising, as an active principle atleast one of the compounds of claim 2, in combination with an acceptablepharmaceutical vehicle.
 9. The pharmaceutical composition according toclaim 8, which is in a form which may be administered orally,parenterally or rectally.
 10. The pharmaceutical composition accordingto claim 8, wherein the dosage of active principle is approximately 0.1to approximately 500 mg/kg/day.
 11. The pharmaceutical compositionaccording to claim 8, which is in a form which may be administeredorally, in a unit dosage of 1 mg to 250 mg of active principle per dose,at the rate of 1 to 4 doses per day.
 12. The pharmaceutical compositionaccording to claim 8, which is in a form which may be administeredparenterally, in a unit dosage of 1 μg to 50 mg of active principle perinjection, at the rate of 1 to 2 injections per day.
 13. A process forpreparation of the compounds of claim 1, wherein Y represents —CONHOH(XV) comprising the following steps: wherein:

step 1 comprises condensing α-hydroxysuccinic acid XII, wherein R₈ is aprotective group compatible with the various elements of the moleculewith an AA-R₃ residue where AA and R₃ are such as defined in claim 21,by a method of coupling used in peptide synthesis, step 2 compriseshydrolyzing the ester XIII obtained in the preceding step intocarboxylic acid XIV with trifluoroacetic acid, step 3 comprises forminghydroxamic acid XV by reacting hydroxylamine, protected O hydroxylamineor diprotected N,O hydroxylamine, with a coupling reagent DCC/HOBT orWSC/HOBT at room temperature in a solvent THF, CH₂Cl₂, or DMF for 1 to24 hours, wherein if R₂=OH, the alcohols are protected prior to saidcoupling; the (di)protected O or N—O hydroxylamines are deprotectedaccording to the nature of the protective group.
 14. The compoundsaccording to claim 1, wherein R₇ is Cl or Br.
 15. The compoundsaccording to claim 1, wherein R₁ is a C₃ to C₁₆ linear or branched, orC₃ to C₆ cyclized alkyl chain comprising O, S, and/or N.
 16. Thecompounds according to claim 1, wherein R₂ is a phenyl substituted byOH.
 17. The compounds according to claim 1, wherein R₂ is a Cl or Br.18. The compounds according to claim 1, wherein R₂ is an amine groupselected from the group consisting of NH₂, NHCOCH₃, CHCOOR₁₀, whereinR₁₀ is a linear or branched C₁ to C₃ alkyl group.
 19. The compoundsaccording to claim 1, wherein R₂ is a methoxycarbonmethyl group or aethoxycarbonmethyl group.
 20. The compounds according to claim 1,wherein AA represents an amino acid in an absolute S configuration andwhich has the formula

wherein R represents: a C₁ to C₄ linear or branched alkyl chain, a—CH₂—Y group in which Y represents a ring of 4 to 6 carbon atoms whichmay be substituted by one of several heteroatoms, said ring beingaromatic or nonaromatic, substituted by one or several —OCH₃, —NO₂, —NH₂groups, or by one or several halogen atoms.
 21. The compounds accordingto claim 20, wherein Y represents a ring of 4 to 6 carbon atoms whichmay be substituted by O, S and/or N, said ring being aromatic ornonaromatic and substituted by one or several halogen atoms.
 22. Thecompounds according to claim 20, wherein Y represents a ring of 4 to 6carbon atoms which may be substituted by one or several heteroatoms,said ring being aromatic or nonaromatic and substituted by Cl, —Br, —Fand/or —I.
 23. The compounds according to claim 1, wherein R₈ is alinear or branched alkyl chain, with 1 to 8 carbon atoms substituted orunsubstituted by O and/or S.
 24. The compounds according to claim 2,wherein R₂ is a methyl or propyl.
 25. The compounds according to claim2, wherein R₂ is a methoxy group.
 26. The pharmaceutical compositionaccording to claim 8, wherein the active principle is in a form whichmay be administered orally and is in a unit dosage of from 10 mg to 250mg of active principle per dose.
 27. The pharmaceutical compositionaccording to claim 8, wherein the dosage of the active principle is from1 to 300 mg/kg/day orally and rectally.
 28. The pharmaceuticalcomposition according to claim 8, wherein the dosage of the activeprinciple is from 0.1 μg/kg/day to 1 mg/kg/day parenterally.
 29. Amethod of preparing a medicine comprising mixing at least one compoundaccording to claim 1 with a pharmaceutically acceptable vehicle.
 30. Amethod of inhibiting the action of a metalloproteinase involved in thebreakdown of the extracellular matrix comprising administering at leastone compound according to claim 1 to a patient in need thereof in anamount sufficient to inhibit said metalloproteinase.
 31. The methodaccording to claim 30, wherein said extracellular matrix is collagenase,gelatinase, or stromelysines.
 32. The method according to claim 30,wherein said patient is an animal or human patient.
 33. A method oftreating a patient suffering from a disease linked to metalloproteinasebreakdown of the extracellular matrix comprising administering to saidpatient an effective amount of at least one compound according to claim1 in an amount sufficient to inhibit the metalloproteinase breakdown ofthe extracellular matrix, wherein said disease is at least one selectedfrom the group consisting of rheumatoid arthritis, osteoarthritis,osteoporosis, corneal ulceration, periodontitis, gingivitis, tumorousinvasions, metastatic proliferation, atherosclerosis, AIDS, chronicinflammatory diseases of the intestine, and neurodegenerative disease.34. The method according to claim 33, wherein said neurodegenerativedisease is Alzheimer's disease or plaque sclerosis.
 35. A method ofinhibiting the release of TNFα from its inactive precursor comprisingadministering at least one compound according to claim 1 to a patient inneed thereof in an amount sufficient to inhibit said release.
 36. Amethod of treating a patient suffering from a disease linked to therelease of TNFα from its inactive precursor comprising administering tosaid patient an effective amount of at least one compound according toclaim 1 in an amount sufficient to inhibit said release, wherein saiddisease is at least one selected from the group consisting of rheumatoidarthritis, Crohn's disease, plaque sclerosis, septic shock, cancer, andcachexia associated with an immunodeficiency.
 37. A method of treating apatient suffering from a disease linked to the production of TNFαcomprising administering to said patient an effective amount of at leastone compound according to claim 1 in an amount sufficient to inhibitsaid production, wherein said disease is at least one selected from thegroup consisting of cancer, psoriasis, eczema, formation of keloids,diabetic retinopathy, atherosclerosis, and inflammatory diseases. 38.Process for preparation of the compounds of formula X such as defined inclaim 1, in which Y represents —CONHOH (also designated hereinafter ascompounds of formula XV), with the exception that R₁ is not aheteroarylalkyl group and that R₂ is not a heteroarylthiomethyl group,characterized in that it comprises: an aldolization reaction from aketo-ester XXVII and alkene XXVIII (in particular in the presence of aLewis acid such as SnCl₄ at −80° C. in a solvent such as CH₂Cl₂ for 5minutes to 2 hours), or from a keto-acid (in the form of a sodium saltor triethylamine) XXX and an alkene XXXI (in particular at roomtemperature between 1 and 10 hours in a THF—H₂O mixture) according tothe following diagram:

 in which: R₁ and R₂ have the meaning indicated hereinabove, R₁₀ is apossibly branched C₁-C₁₂ alkyl, a benzyl or an optically pure compoundsuch as mandelic acid esterified with a linear or branched C₁-C₃ alkyl,or a benzyl, R₁₁ is a linear or branched C₁-C₃ alkyl, or a chlorine, R₁₂is sodium or triethylamine, R₁₃ is hydrogen, a linear or branched C₁-C₃alkyl; R₁₃ also may represent a chain forming a ring with the boron atomsuch as, for example, di-isopropyltartrate, the reactions arediastereoselective and lead to stereochemistry derivatives XVI if thedouble bond is of Z geometry for the compounds XXVIII and E geometry forthe compounds XXXI, the obtaining of the aforementioned compounds offormula XV then being accomplished according to the following reactiondiagram:

 in which: steps 4 and 6 are performed as in steps 1 and 3 of thereaction diagram of claim 17 respectively, and starting from compoundsXVI and XIV, which leads to the compounds of formula XVII and XVrespectively, step 5 consists in oxidizing the double ethylene bond ofthe compound of formula XVII into acid, in particular by ozonolysis (forexample at −60° C. in CH₂Cl₂ until obtaining a steady blue color), thenoxidation (in particular at room temperature with NaClO₂ and NaH₂PO₄ intBuOH—H₂O for 15 hours) or directly by KMnO₄/NalO₄ (in particular atroom temperature in a tBuOH—H₂O mixture for 1 to 10 hours), which leadsto the compound of formula XIV.
 39. Process for preparation of thecompounds of formula X such as defined in claim 1, in which Yrepresents: —SH, or a group with the formula

 or a group with the formula

in which R₄ and R₅ are such as defined in claim 1, said process beingperformed according to the following diagram:

in which: step 44 consists in opening an epoxide of formula L in whichR₁ and R₂ are such as defined in claim 1, and R₉ is a carboxylic acidprotective group, in particular a benzyl remainder sensitive tocatalytic hydrogenolysis, this opening of the epoxide L beingaccomplished with a nucleophile, for example a thiol protected by an R₁₈group compatible with R₉, for example a benzyl in methanol for 1 hour at60° C., step 47 consists in deprotecting the ester LI, for example withtrifluoroacetic acid as before, step 48 is identical to step 1 of thereaction diagram of claim 17, and performed starting from compound LIIobtained in the preceding step, step 49 consists in deprotecting thesulfur, for example with sodium in liquid ammonia, in particular at −60°C. for 5 to 15 minutes, step 45 consists in opening the epoxide L withprotected hydroxylamine such as defined in step 3 of the processaccording to claim 17 (with, for example, R₁₉=benzyl or THP), step 50consists in deprotecting the ester LV by a method compatible with R₁₉,step 51 is identical to step 48, and performed starting from thecompound LVI obtained in the preceding step, step 52 consists inreacting the hydroxylamine LVII with formic acid and acetic anhydride,in particular at a temperature of at least 100° C. for 1 to 15 hours,step 53 consists in cleaving R₁₉ on the compound LVIII with, forexample, H₂ Pd/C or HCl 1N depending on the structure of R₁₉ as before,step 46 consists in opening an epoxide L with hypophosphorous acid withthe formula H₃PO₂, then esterification with a coupling agent such as DCCand an R₄OH group in which R₄ is such as defined in claim 1, in thepresence, for example, of trimethylorthoformate and tetramethylguanidineat room temperature for 5 hours, step 54 consists in treating thecompound LIX with a compound with the formula

 (prepared according to the methods described in the literature) inwhich R₆ and R₇ are such as defined hereinabove, in particular in CH₂Cl₂in the presence of bis trimethylsilyl acetamide at room temperature for5 hours, which leads to the obtaining of the compound of formula LX inwhich R₅ represents:

step 55 consists in cleaving the R₉ ester by a method compatible with R₄as previously, step 56 is identical to step 48, and performed startingfrom the compound LXI obtained in the preceding step, step 57 consistsin cleaving the R₄ group of the compound LXII obtained in the precedingstep, for example with the aid of Nal in acetone under reflux for 15hours.